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annotated research ‘Annotated research methods in Caribbean Research’ (Cook Bastick, 2010) The editors are issuing a call for papers for inclusion in the printed publication ‘Annotated research methods in Caribbean Research'. For more information, see below. This book of papers is intended for essay gill Caribbean researchers who need to expand their repertoire of research methods in order to release their ideas for rigorous published research. This is an essay theory innovative approach to enhancing research relevant to the Caribbean and its Diaspora. The editors are of the view that an extended repertoire of method options is essay gill a timely conceptual tool for chris hoover thesis promoting rigorous development of on typography gill Caribbean research and for enabling the rich diverse research potential of our Caribbean researchers. Way To. To this end we invite the original authors of Caribbean research to submit for this publication their ALREADY PUBLISHED PEER-REVIEWED PAPERS with additional personal annotations of the research methods they have used.
Annotated methods papers will be published under the following sections and categories, according to the main methods used: Quantitative various categories Qualitative various categories Mixed methods various categories. Innovative methods are particularly welcome. The book will be indexed under the essay on typography gill, areas of research covered by Caribbean scholars and practitioners with disciplinary affiliations in health, education, psychology, communications, languages, development, management, social work, Caribbean identity, cultural studies and other relevant areas. Format of submitted papers. Your original published paper may be annotated in one of two formats (i) Embedded : Method annotations can be embedded at appropriate points within the original paper, using a different font or (ii) Endnotes : Numbers (similar to mla format, footnote numbers) can be used at the relevant sections in the original paper. Essay On Typography Gill. These numbers will then refer to essay, endnotes; that is, all the annotations corresponding to the numbers are placed at the end of the original published paper rather than as footnotes that are placed at essay gill, the bottom of each page. However, whichever format authors prefer, it is transport technology necessary to on typography gill, begin with an Abstract of quoting their method annotations followed by an Introduction relating their chosen methods to their Caribbean content. The original annotated paper would then follow and conclude with a Discussion of the practical/theoretical advantages and on typography, disadvantages, the achieved/thwarted intentions of the author and personal advice to the readers on best way to essay improvements that the on typography, author would, in 20:20 hindsight vision, now make if replicating the research.
The annotated parts of the final paper can be approximately 3,000 to 5,000 words in addition to transport, the word length of the original paper. Style is essay on typography gill APA (5th Edition). Sports Prompts. Manuscripts should be sent as email attachments. Diagrams/Figures should be embedded in your manuscript using Windows metafile format or Enhanced metafile format, whichever enables you to enlarge without loss of on typography gill picture quality (to do this copy and use the ‘paste special’ choices). Authors are responsible for obtaining the necessary copyright releases, if any, from the publishers of their original papers. These releases must be submitted with the annotated paper. Essay Transport Technology. At the initial stage, a draft abstract can be submitted without copyright releases.
The editors promote open-access to research. To this end, the book will be published under Creative Commons copyright* so that readers may use the work without permission, providing they give full credit to essay gill, the authors, do not use the work for commercial advantage and pass this access on essay with their use of the work. Authors similarly retain full rights to their published work. Please note that under most Copyright Acts, the essay on typography, author of an article may retain the thesis, copyright to an article accepted for journal publication. On Typography. However, some closed-access journals demand the transfer of copyright as a condition of publishing your article. Even though ours is a non-profit publication for educational purposes, if you have signed away copyright of oil characterization your paper, as the author, you are responsible for obtaining written consent from the original copyright holder for gill Attribution-Non-commercial-Share Alike Creative Commons world-wide usage*, submitting all original signed permissions to us when you submit your manuscript, and paying any usage fees to your copyright holder. If you have signed away copyright in your work you are advised to enquire with your copyright holder before submitting your intention to publish in this book.
We will email a ‘Permission request letter’ to you, should you need one. To download a Copyright Request Form, and the Guidelines for structuring your annotated paper, please enter below your name and oil characterization thesis, the email address of where the form and guidelines should be sent : Six-step process to publication. Essay. Step 1: Choose one of your peer-reviewed published papers on Caribbean research for which you would like to annotate your methodology. Step 2: Check if you have signed away your copyright. If so, then download the for essays quoting, copyright request form and on typography, send it to the copyright holder for permission to republish your paper in our book. Thesis. Step 3: Write a draft abstract (about 250-300 words) saying very briefly what aspect of Caribbean research your paper covers, what methods you used and why you think they are particularly suitable for this aspect of Caribbean research. Step 4: Email your ‘notice of intent’ to one of the editors. This will include these three attachments: (i) Your draft abstract, with a statement of the section and category in which you would tentatively place the essay on typography, research method(s) you will be annotating, (ii) the essay technology, citation of your published paper with a copy of that paper, and essay on typography, (iii) a statement saying you hold the copyright or that you are seeking the permission to republish from the copyright holder. YOU CAN DO IT NOW. This must be completed by the first deadline of Jan 14th 2008.
Within two-weeks you will then hear from the editors if we can accept your submission. Step 5: Once your submission has been accepted, you should prepare your annotated paper for review as outlined in mla format for essays, the ‘Format of submitted papers’ from the Call for papers on www.AnnotatedResearch.com and email it to the editors, with the completed copyright release if required, and brief ‘bio-pic’ that introduces you to the readers. This must be completed by gill the second deadline of Feb 29th 2008. Step 6: Where necessary, the sports essay prompts, editors will identify a reviewer to work with you in fine tuning the annotations to suit the readership of the book. Your final annotated paper must be ready for publication by essay on typography the third deadline of May 28th 2009. The editors will then work with the essay transport technology, publishers to have the book published by April 2010. Notice of intent: YOU CAN DO IT NOW, and up to the very last day of January, Thurs Jan 31st 2008: indicate your intention to gill, submit your annotated methods paper by sending a draft abstract of your intended annotations, an APA citation to your original paper, a copy of your origina l paper and an indication of the section and category in which you would tentatively place the research method(s) you will be annotating.
Paper for review : Feb 29th 2009: draft annotated methods paper for review, with any necessary copyright releases. Paper for publication : March 28th 2010 final annotated methods paper for publication. Please send your ‘Notice of intent’ (see above) to:
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Smalls-Mantey, Sample F31 Application and on typography Summary Statement. We are truly indebted to the grantee who allowed us to mla format, post this outstanding F31 sample application to help the next generation of investigators write applications. The F31 sample application below doesn’t reflect the essay, complex layouts of the actual SF 424 PDF forms. Instead, we’ve provided a list of key fields from those forms and the applicant’s responses. In addition, we redacted some applicant-specific information such as contact information, publications in press, and budget specifics. The text of quoting, this application is copyrighted. You may use it only for essay on typography gill nonprofit educational purposes provided the document remains unchanged and the PI, the grantee organization, and NIAID are credited. Questions?
Contact firstname.lastname@example.org. We also offer other outstanding Sample Applications. [Our Web version of this sample application can't reproduce the best write an opinion, layout of the actual PDF forms. Here is the on typography gill, summary that eRA Commons generated at the start of the oil characterization thesis, sample app, which reflects many key fields from on typography, that form.] Project/Performance Site Locations.
[Our Web version of this sample application can't reproduce the layout of the actual PDF forms. Performance sites are in New York, Maryland, and the United Kingdom.] Research Related Other Project Information. [Our Web version of this sample application can't reproduce the layout of the actual PDF forms. Here are the essay, key fields from that form and essay gill the applicant's responses.] Understanding the sports, basis of an immune response that controls infection or provides sterilizing immunity remains a major goal in the search for effective vaccines or immunotherapies for essay HIV. Essay! Antibodies (Abs) induced by candidate vaccines to the surface envelope glycoprotein have not neutralized a broad array of primary virus isolates. For this reason, eliciting a cytotoxic cellular response has been the primary goal in most recent vaccine trials.
However, this approach has not been successful in containing viral replication in essay on typography, vaccinees that have become HIV-infected. Antibody-dependant cellular cytotoxicity (ADCC) has been shown to mediate sterilizing immunity against thesis, challenge with pathogenic simian immunodeficiency virus [Hessel 2007]. In ADCC, Fc-bearing Abs bind viral epitopes coating an infected CD4+ target T cell and an Fc receptor bearing effector, most commonly natural killer cells (NKs), bind the Ab and use perforin to deliver granzymes which induce apoptosis in the target. We want to study ADCC in infected patients to understand the magnitude and characteristics of the best responses achieved by natural infection. First, we will compare ADCC mediated by essay on typography gill the sera of essay on the of evolution, a cohort of patients using a granzyme B cytotoxicity assay developed in our lab. Based on these findings, we will select the on typography, sera of patients with the most ADCC, generate monoclonal Abs (mAbs), and characterize the mAbs based on oil characterization thesis epitope specificity, affinity, potency, breadth, IgG isotype, and Fc type.
We will also evaluate whether ADCC is disparate from classical neutralization. Essay Gill! Finally, we will use microscopy to examine the synapse between effectors, Abs, and targets. The outcome of this research will provide insight into the characteristics of Abs that mediate ADCC that are likely important goals in the design of HIV vaccines or immunotherapies. Hypothesis: Antibody-dependent cellular cytotoxicity (ADCC) is a function that has been shown to mediate protection from lentiviral infection. We hypothesize that variations in ADCC activity of sera are dictated by the amount, specificity, and subclass of HIV-specific antibodies. Aim 1 : Characterize the sports, potency of sera of on typography, HIV-infected individuals in ADCC. Aim 2 : Characterize the theory of evolution, specificity and breadth of antibodies with ADCC activity. Aim 3 : Characterize the structure and essay on typography function of the best way to, target-effector synapse using both fixed and essay live cell laser scanning confocal microscopy (LSCM), transmission electron microscopy (TEM) and essay transport cryo-electron microscopy (cryo-EM) and tomography. Understanding the role of antibody-dependent cellular cytotoxicity (ADCC) in HIV could provide important insights for induction of this activity through vaccination.
This project seeks to characterize the Abs that mediate ADCC and image the functional synapse formed by cellular components involved in ADCC with the goal of defining new goals for the development of HIV vaccines and therapeutics. Gallo RC, Salahuddin SZ, Popovic M. Frequent detection and isolation of cytopathic retroviruses (HTLV-III) from patients with AIDS and at risk for AIDS. Science. 1984; 224: 500-3. Barre-Sinoussi F, Chermann JC, Rey F, Nugeyre MT, Chamaret S, Gruest J, et al. Essay! Isolation of a T-lymphotropic retrovirus from best an opinion, a patient at on typography gill, risk for acquired immune deficiency syndrome (AIDS).
Science. 1983; 220(4599): 868-71. Organization WH. Global summary of the AIDS epidemic, December 2007. 2007 [cited 2009 September 18, 2009]; Hessell AJ, Hangartner L, Hunter M, Havenith CE, Beurskens FJ, Bakker JM, et al. Fc receptor but not complement binding is important in antibody protection against HIV. Nature. 2007; 449(7158): 101-4. www.hivresearh.org. RV144 Phase III HIV Vaccine Trial Press Release. 2009. Chung A, Rollman E, Johansson S, Kent SJ, Stratov I. The utility of ADCC responses in HIV infection.
Current HIV research. 2008; 6(6): 515-9. Kohl S, Starr SE, oleske JM, Shore SL, Ashman RB, Nahmias AJ. Human monocytemacrophage- mediated antibody-dependent cytotoxicity to herpes simplex virus-infected cells. J Immunol. Hoover! 1977; 118(3): 729-35. Forthal DN, Landucci G. In vitro reduction of virus infectivity by antibody-dependent cellmediated immunity. On Typography! Journal of immunological methods. 1998; 220(1-2): 129-38.
Stratov I, Chung A, Kent SJ. Essay Theory Of Evolution! Robust NK cell-mediated human immunodeficiency virus (HIV)-specific antibody-dependent responses in HIV-infected subjects. J Virol. 2008; 82(11): 5450-9. Gill! Gomez-Roman VR, Florese RH, Patterson LJ, Peng B, Venzon D, Aldrich K, et al.
A simplified method for the rapid fluorometric assessment of antibody-dependent cell-mediated cytotoxicity. Journal of immunological methods. Thesis! 2006; 308(1-2): 53-67. Migueles SA, Osborne CM, Royce C, Compton AA, Joshi RP, Weeks KA, et al. Lytic granule loading of CD8+ T cells is essay on typography gill, required for thesis HIV-infected cell elimination associated with immune control. Immunity. Essay On Typography! 2008; 29(6): 1009-21. Vago L, Perna SK, Zanussi M, Mazzi B, Barlassina C, Stanghellini MT, et al. Essay Prompts! Loss of mismatched HLA in leukemia after stem-cell transplantation.
N Engl J Med. 2009; 361(5): 478- 88. Ziegner U, Campbell D, Weinhold K, Frank I, Rutstein R, Starr SE. Deficient antibodydependent cellular cytotoxicity against human immunodeficiency virus (HIV)-expressing target cells in perinatal HIV infection. Clin Diagn Lab Immunol. 1999; 6(5): 718-24. Brunetta E, Fogli M, Varchetta S, Bozzo L, Hudspeth KL, Marcenaro E, et al. The decreased expression of Siglec-7 represents an early marker of dysfunctional natural killer cell subsets associated with high levels of HIV-1 viremia. Blood.
2009. Ljunggren HG, Ohlen C, Hoglund P, Yamasaki T, Klein G, Karre K. Afferent and efferent cellular interactions in natural resistance directed against essay gill, MHC class I deficient tumor grafts. J Immunol. Of Evolution! 1988; 140(2): 671-8. De Maria A, Ferraris A, Guastella M, Pilia S, Cantoni C, Polero L, et al. Expression of essay on typography gill, HLA class I-specific inhibitory natural killer cell receptors in HIV-specific cytolytic T lymphocytes: impairment of specific cytolytic functions. Best Way To Write Essay! Proceedings of the National Academy of Sciences of the United States of America.
1997; 94(19): 10285-8. Ahmad R, Sindhu ST, Toma E, Morisset R, Vincelette J, Menezes J, et al. Evidence for essay on typography a correlation between antibody-dependent cellular cytotoxicity-mediating anti-HIV-1 antibodies and prognostic predictors of HIV infection. J Clin Immunol. 2001; 21(3): 227-33. Lambotte O, Ferrari G, Moog C, Yates NL, Liao HX, Parks RJ, et al. Sports Essay Prompts! Heterogeneous neutralizing antibody and antibody-dependent cell cytotoxicity responses in HIV-1 elite controllers.
AIDS. 2009; 23(8): 897-906. Dalgleish A, Sinclair A, Steel M, Beatson D, Ludlam C, Habeshaw J. Failure of ADCC to essay, predict HIV-associated disease progression or outcome in a haemophiliac cohort. Oil Characterization! Clin Exp Immunol. 1990; 81(1): 5-10.
Koup RA, Sullivan JL, Levine PH, Brewster F, Mahr A, Mazzara G, et al. Antigenic specificity of gill, antibody-dependent cell-mediated cytotoxicity directed against human immunodeficiency virus in antibody-positive sera. Essay On The! J Virol. 1989; 63(2): 584-90. Doria-Rose NA, Connors M. Antibody-secreting B cells in HIV infection. Curr Opin HIV AIDS. 2009; 4(5): 426-30.
Burke B, Barnett SW. Broadening our view of protective antibody responses against HIV. Current HIV research. 2007; 5(6): 625-41. Sattentau Q. Avoiding the void: cell-to-cell spread of human viruses.
Nat Rev Microbiol. Essay! 2008; 6(11): 815-26. Chamow SM, Zhang DZ, Tan XY, Mhatre SM, Marsters SA, Peers DH, et al. A humanized, bispecific immunoadhesin-antibody that retargets CD3+ effectors to thesis, kill HIV-1- infected cells. J Immunol. On Typography! 1994; 153(9): 4268-80. Jolly C, Kashefi K, Hollinshead M, Sattentau QJ. HIV-1 cell to cell transfer across an thesis, Env-induced, actin-dependent synapse. J Exp Med. 2004; 199(2): 283-93. Groot F, Welsch S, Sattentau QJ.
Efficient HIV-1 transmission from macrophages to T cells across transient virological synapses. Blood. On Typography Gill! 2008; 111(9): 4660-3. Martin N, Welsch S, Jolly C, Briggs JA, Vaux D, Sattentau QJ. Virological synapsemediated spread of human immunodeficiency virus type 1 between T cells is sensitive to entry inhibition. J Virol. 2010; 84(7): 3516-27. Intellectual Support : My mentors Mark Connors and essay on the theory of evolution Quentin Sattentau are leading experts in their field and gill have mentored numerous people that have gone on to successful careers in science. Mla Format For Essays! This dual mentorship will teach me to aggressively approach science from two distinctive perspectives in my graduate career, something most PhD scientist don’t experience until their first post-doctoral experience.
While at NIH I will have received extensive support of staff from Richard Siegel, Director of the NIH MD/PhD partnership program and Michael Lenardo, Director the NIH/Oxford/Cambridge Scholars Program. While at essay on typography gill, Oxford I will have the essay prompts, guidance of Lucinda Risius, Managing Director of the Graduate Pathology Program and I will also be assigned a departmental advisor, outside of my lab, with whom I can discuss my research Progress. Educational Resources : Through the FAES at NIH I have access to courses to improve lab specific skills and foundation knowledge. I have already participated a Vaccine Development course. At Oxford I will take seminars at the Centre for Excellence in Teaching and Learning for training in teaching skills.
I will also take courses pertinent to my research offered at the Dunn School of essay gill, Pathology. Both the NIH and Oxford have libraries and IT staff that can aid me in scholarly research. Physical Resources : Both labs have sufficient funding to provided me any reagent/equipement need for transport technology the proposed research. Lab Space – I am provided with a desk and personal desktop computer. Bench space is shared within the lab. Safety – In order to essay gill, ensure our safety from hazardous blood samples we work under laminar flow hoods in a P2 facility with P3 practices. Samples – We receive blood samples from patients enrolled within our protocol. We also have access to the NIH blood blank for normal human blood samples. As provided by Quentin Sattentau: “Oxford University is thesis, held in high international regard for its strong teaching and research background and its unique and personal teaching style.
Within the University the Department of Pathology has an excellent reputation for graduate teaching, and on typography gill has achieved 100% success in DPhil graduation within 4 years for the past 3 years. The Department has a strong graduate support structure, in best essay, which each student has a principal supervisor, the possibility of a cosupervisor, a departmental advisor, and a college advisor. The supervisor(s) will generally meet with the student multiple times a term, and each of the non-supervisory mentors will meet with the candidate approximately once a term and essay on typography will file reports on thesis the candidate’s progress. The candidate will be expected to essay on typography, attend a series of high profile external seminars that take place once a week in term time, and a regular series of internal seminars from leading and more junior University and essay prompts Departmental investigators and essay on typography students. Adjoa will attend and participate in the weekly lab journal club, in which scientific papers are taken apart critically by members of the group, and oil characterization thesis a weekly laboratory meeting in which each lab member presents their ongoing research. Adjoa will also be encouraged to chat to other members of the department during tea lunch and coffee breaks, a longstanding Oxford institution! The Department has a fully equipped quiet library for literature research and other reading and writing. Adjoa will have her own writing up space close to her work bench, equipped with a portable computer and if required, an external screen.
Adjoa will have full access to all core facilities, including the imaging suite with multiple light microscopes, a flow cytometry and essay gill sorting suite, an electron microscopy facility with full technical and scientific support. Mla Format Quoting! We also have strong connections with the biochemistry and chemistry departments and the Weatherall Institute of Molecular Medicine at the John Radcliffe Hospital for obtaining clinical samples and expertise.” BD FACSAria IIu. BD Pathways 855. VICTOR Light luminescence plate reader. AutoMACS Pro Separator. AutoMACS Classic Separator.
Containment II/III Laboratory with 4 tissue culture hoods. Tissue culture. The laboratory has a new tissue culture facility comprised of 8 double hoods and associated incubators and ancillary equipment that is shared with three other virology groups. Virology. Essay On Typography! The department has a large, fully-equipped containment-III laboratory shared between two groups, with three tissue culture hoods and associated incubators and ancillary equipment, in which HIV-1 is grown and used to infect cells. Flow cytometry. The department has a core flow cytometry facility with three BD analyzers (2 x FACScalibur 2 x LSR2) and two sorters. Molecular biology. The Sattentau laboratory has all facilities for molecular biology carried out under containment levels –I, -II and –III. Light microscopy.
The department has 5 confocal microscopes of thesis, varying sophistication and resolution, two of which are set up for live cell immunofluorescence work. The department has three recently acquired high-speed CCD cameras and associated microscopes for live cell immunofluorescence analysis. The Sattentau laboratory has a Zeiss Pascal confocal microscope and essay on typography gill a new CCD camera set up with a Zeiss Axiovert 200 microscope for fixed and live cell work in the containment-III laboratory, allowing real time analysis of live, infected cells. Electron microscopy. The department has two transmission electron microscopes and a scanning electron microscope, all of which are maintained and run as a central service facility by a skilled dedicated operator. Essay On The Theory Of Evolution! Our collaborator in Oxford (Kay Grunewald) is located at the University Churchill campus, which is 4 miles from the on typography, Pathology Department. He maintains and uses 3 microscopes set up for cryo-tomography as follows.
300keV FEG cryo-electron microscope FEI TF30 Helium Polara with GIF2002 imaging filter (2 x 2k CCD) 300keV FEG cryo-electron microscope FEI TF30 with 4 x 4kCCD 120keV cryo-electron microscope FEI T12 with 4 x 4k CCD. Research Related Senior/Key Person Profile. [Our Web version of this sample application can't reproduce the layout of the actual PDF forms. See below for the biographical sketch attachments.] My interest in studying infectious diseases stems from the 2001 anthrax scare in the Washington D.C. area. I already had a strong interest in science but through my participation in the Gene Search Science Project at Catholic University the following summer I came to understand how medical research impacts treatment for routine illnesses and also plays a role in national defense. Since middle school I have been preparing for a career in on the of evolution, science. At Eleanor Roosevelt High School I was enrolled in the rigorous Science and essay on typography gill Technology Program during which time in participated in oil characterization, the Gene Search Project and researched biochemical pathways that influence sickle cell anemia with Dr. William Winters at Howard University Hospital. This experience taught me how research and medicine are intertwined and inspired me to become a physicianscientist. On Typography! At UMBC I was a member of the Meyerhoff Scholars Program and on the theory of evolution an HHMI Undergraduate Research Scholar where numerous lab experiences, seminars, rigorous classes, and multiple opportunities for essay scientific presentations prepared me for a career as a scientist.
My lab skills and understanding of a research career increased exponentially during my four years in Michael Summers’ lab. Transport Technology! Not only did I learn about protein purification, gel electrophoresis, NMR, and ITC while studying MuLV RNA packaging, but more importantly of the time and essay on typography discipline necessary for excellent research and the internal drive that keeps one focused. Working with Dr. Nikola Pavletich I learned the value of immediately organizing my data so that others could analyze my results quickly. In the lab of Tom Cech I had the thesis, responsibility of designing a new protocol to study telomerase assembly, a skill that has enabled me to design a novel assay to essay gill, investigate AIDS specific questions in my graduate studies. The medical education I’ve received at Columbia has given me a unique perspective with which I approach scientific questions. Under the tutelage of Mark Connors and Quentin Sattentau, leaders in essay on the of evolution, their fields on nonprogressive HIV infection and viral transmission, my understanding of gill, infectious disease and my capacity to pursue further research in these fields will be greatly enhanced. Essay! In addition, the on typography, courses, medical preceptorships, presentations, teaching, and career development activities I am undertaking in the forthcoming years make me an excellent P.I. for this grant.
Academic and Professional Honors. 2003 National Urban League Scholarship 2003 UMBC Meyerhoff Scholar 2003 UMBC Honors College 2003 HHMI Undergraduate Research Scholar 2004 NAD Distinguished Youth Award 2005 UMBC Minority Access to way to write an opinion, Research Careers Scholar 2005 Abby Rockefeller Mauze Award Charitable Trust Scholar for essay gill Outstanding Achievement 2006 Goldwater Scholar 2007 UMBC Top Senior Biochemistry Major 2007 Graduated Sum Cum Laude from UMBC 2007 NIH/Oxford/Cambridge Scholar 2010 International AIDS Vaccine Conference Scholarship 2011 Keystone Symposia Underrepresented Minority Scholarship. 2004 Omicron Delta Kappa Leadership Honor Society 2004 Golden Key International Honor Society, Sophomore/Alumni Director 2007 Phi Beta Kappa. Dey A, York D, Smalls-Mantey A, Summers MF. Sports Prompts! 2005. Composition and sequence-dependent binding of RNA to gill, the nucleocapsid protein of sports essay, Moloney murine leukemia virus. Biochemistry.
44(10), 3735-3744. Miyazaki Y, Irobalieva RN, Tolbert BS, Smalls-Mantey A, Iyalla K, Loeliger K, D'Souza V, Khant H, Schmid MF, Garcia EL, Telesnitsky A, Chiu W, Summers MF. 2010. Structure of a conserved retroviral RNA packaging element by NMR spectroscopy and cryo-electron tomography. J Mol Biol.
404(5), 751-772. Smalls-Mantey A, Winter W. 2003. Gill! Biosynthesis of 2,3-diphosphoglycerate in best way to write, Erythrocytes. Essay On Typography! Poster presentation at ERHS Science Fair and Prince Georges County Science Fair. March 2003. Oil Characterization Thesis! Smalls-Mantey A, Dey, A, Phillip C, Summers MF.
2004. Characterization of High-Affinity Nucleocapsid Protein Binding Sites Within the Moloney Murine Leukemia Virus RNA Packaging Signal. Poster presented at essay gill, 2004 Annual Biomedical Research Conference for Minority Students, November 2004. Smalls-Mantey A, Min J, Pavletich N. 2005. Structural and theory of evolution Biochemical Studies of the Yeast Rad4-Rad23 Complex Bound to DNA. Poster presented at Tri-Institutional Gateways to on typography gill, the Laboratory Program Symposium and chris hoover thesis 2005 Leadership Alliance.
August 2005. Smalls-Mantey A, Miyazaki Y, Summers MF. 2007. Dimerization of Moloney Murine Leukemia Virus Nucleocapsid Protein Binding Sites Facilitates Binding. Poster presented at on typography gill, 2007 UMBC Undergraduate Research and Creative Achievement Day. April 2007. Smalls-Mantey A, Klein R, Doria-Rose N, Laub L, Rood J, Migueles S, Sattenatu. 2010. HIV-Specific Antibodies Mediate Rapid Antibody-Dependent Cellular Cytotoxicity Against Primary HIV-Infected CD4+ T Cells.
Poster presented at 2010 Keystone HIV Vaccines (X5) Meeting. On The! March 2010. On Typography Gill! Smalls-Mantey A, Klein R, Doria-Rose N, Laub L, Rood J, Migueles S, Sattenatu. 2010. HIV-Specific Antibodies Mediate Rapid Antibody-Dependent Cellular Cytotoxicity Against Primary HIV-Infected CD4+ T Cells. Poster presented at 2010 Oxford Pathology Department Graduate Student Symposium.
June 2010. Smalls-Mantey A, Klein R, Doria-Rose N, Laub L, Rood J, Migueles S, Sattenatu. 2010. HIV-Specific Antibodies Mediate Rapid Antibody-Dependent Cellular Cytotoxicity Against Primary HIV-Infected CD4+ T Cells. Poster presented at 2010 NIH/Oxford/Cambridge Scholars Research Colloquium. Way To An Opinion! Smalls-Mantey A, Klein R, Doria-Rose N, Laub L, Rood J, Migueles S, Sattenatu. 2010. HIV-Specific Antibodies Mediate Rapid Antibody-Dependent Cellular Cytotoxicity Against Primary HIV-Infected CD4+ T Cells.
Poster presented at 2010 International AIDS Vaccine Conference. On Typography! September 2010. Smalls-Mantey A. 2010. Essay Prompts! Antibody-Dependent Cellular Cytotoxicity and the Control of essay on typography gill, HIV. Talk given at for essays, Frontiers in Biomedicine NIH/Oxford/Cambridge Scholars Program student presentation series. November 2010. [Outside research support redacted from sample.] A. Personal Statement Regarding Project. Currently HIV-specific ADCC is considered one of the very top priorities in the field of HIV-specific immunity. Essay! Our laboratory has assembled the best techniques and reagents in the field to carry out this project.
We have extensive experience in HIV-specific antibodies and cytotoxicity. Oil Characterization! In addition we have a long track record of training students to prepare them for a carreer in biomedical research. 1985 - 1986 Intern in Pediatrics, The Boston Floating Hospital, New England Medical Center, Boston, Massachusetts 1986 - 1988 Resident in essay on typography, Pediatrics, The Boston Floating Hospital, New England Medical Center, Boston, Massachusetts 1988 – 1989 Chief Resident in Pediatrics, The Boston Floating Hospital, New England Medical Center, Boston, Massachusetts July 1989 - October 1989 Medical Staff Fellow, Respiratory Viruses Section, Laboratory of Infectious Diseases, National Institute of way to write, Allergy and on typography Infectious Diseases, National Institutes of Health, Bethesda, MD October 1989 - June 1993 U.S. Public Health Service, Research Associate (C.O.), Respiratory Viruses Section, Laboratory of Infectious Diseases, National Institute of mla format for essays, Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD July 1993 - June 1994 U.S. Public Health Service, Clinical Associate, Clinical Center, National Institute of essay on typography, Allergy and Infectious Diseases, National Institutes of thesis, Health, Bethesda, MD January 1994 - June 1994 Clinical Fellow in Infectious Diseases, The Childrens Hospital of Philadelphia, Philadelphia, PA. June 1994 - December 2004 U.S.
Public Health Service, Medical Officer, National Institute of Allergy and Infectious Diseases, Clinical and Molecular Retrovirology Section, Laboratory of Immunoregulation, National Institutes of essay, Health, Bethesda, MD December 2007- Present Chief, HIV-Specific Immunity Section, Laboratory of thesis, Immunoregulation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD. Recent Honors and essay on typography gill Editorial and Review Board Activities. 2009 - U.S. Public Health Service Outstanding Service Medal 2009 - present Vaccine Production Development Team Scientific Advisory Board, University of Pennsylvania 2009 - present HIV Enterprise Host Genetics and HIV Diversity Working Group 2010 - present Editorial Board, Journal of Virology. C. Peer-Reviewed Publications. Migueles SA, Sabbaghian MS, Shupert WL, Bettinotti MP, Marincola FM, Schwartz D, Sullivan J, and Connors M. HLA B*5701 is essay transport, highly associated with restriction of virus replication in a subgroup of HIV infected long term nonprogressors. PNAS 2000; 97(6): 2709-2714. Migueles SA, Laborico AC, Shupert WL, Sabbaghian MS, Rabin R, Hallahan C, Van Baarle D, Kostense S, Miedema F, McLaughlin M, Ehler L, Metcalf J, Liu S, and Connors M. HIV-specific CD8+ T cell proliferation is coupled to perforin expression and is maintained in nonprogressors. Nature Immunol 2002; 3(11); 1061-1068. Migueles SA, Laborico AC, Imamichi H, Shupert WL, Royce C, McLaughlin M, Ehler L, Metcalf J, Liu S, Hallahan CW, and Connors M. Essay! The differential ability of HLA B*5701+ long-term nonprogressors and progressors to way to write, restrict HIV replication is not caused by on typography gill loss of recognition of autologous viral gag sequences.
J Virol 2003; 77(12); 6889-6898. Iyasere C, Tilton JC, Johnson AJ, Younes S, Yassine-Diab B, Sekaly RP, Kwok WW, Migueles SA, Laborico AC, Shupert WL, Hallahan CW, Davey RT Jr., Dybul M, Vogel S, Metcalf J, and Connors M. Diminished proliferation of for essays, HIV-specific CD4+ T Cells is associated with diminished IL-2 production and recovered by on typography exogenous IL-2. J Virol 2003; 77(20); 10900-10909. Tilton JC, Luskin MR, Johnson AJ, Manion M, Hallahan CW, Metcalf JA, McLaughlin M, Davey RT Jr., and Connors M. Changes in essay technology, paracrine IL-2 requirement, CCR7 expression, frequency, and cytokine secretion, of essay on typography, HIV-specific CD4+ T cells are a consequence of chris hoover thesis, antigen load. Essay! J Virol 2007; 81;2713-2725. Migueles SA, Osborne CM, Royce C, Compton AA, Joshi RP, Weeks KA, Rood JE, Berkley AM, Sacha JB, Cogliano-Shutta NA, Lloyd M, Roby G, Kwan R, McLaughlin M, Stallings S, Rehm C, O’Shea MA, Mican J, Packard B, Komoriya A, Palmer S, Wiegand AP, Maldarelli F, Coffin JM, Mellors JW, Hallahan CW, Follman DA and Connors M. Lytic Granule Loading of chris hoover thesis, CD8+ T-Cells is Required for essay gill HIV-Infected Cell Elimination Associated with Immune Control. Immunity 2008; 29 (6): 1009-1021. Migueles SA, Weeks KA, Nou E, Berkley AM, Rood JE, Osborne CM, Hallahan CW, Cogliano-Shutta NA, Metcalf JA, McLaughlin M, Kwan R, Mican JM, Davey RT Jr, Connors M. Thesis! Defective human immunodeficiency virus-specific CD8+ T-cell polyfunctionality, proliferation, and cytotoxicity are not restored by antiretroviral therapy. J Virol 2009 83(22): 11876-11889. Doria-Rose NA, Klein R, Daniels M, O'Dell S, Nason M, Lapedes A, Bhattacharya T, Migueles S, Wyatt RT, Korber BT, Mascola JR, Connors M. Breadth of HIV-Specific neutralizing activity in sera: clustering analysis and association with clinical variables. J Virol 2009 84(3): 1631-1636.
Doria-Rose NA and Connors M. Antibody-secreting B cells in HIV infection. Curr Opin HIV AIDS 2009 4:426-430 Wu X, Yang ZY, Li Y, Hogerkorp CM, Schief WR, Seaman MS, Zhou T, Schmidt SD, Wu L, Xu L, Longo NS, McKee K, O'Dell S, Louder MK, Wycuff DL, Feng Y, Nason M, Doria-Rose N, Connors M, Kwong PD, Roederer M, Wyatt RT, Nabel GJ and Mascola JR. Rational design of envelope identifies broadly neutralizing human monoclonal antibodies to HIV-1. On Typography Gill! Science 2010 329:856-861. Mla Format! Migueles SA and essay on typography gill Connors M. Long-term nonprogressive disease among untreated HIV-infected individuals: clinical implications of chris thesis, understanding immune control of HIV. JAMA 2010 304:194-201. [Redacted publication in press]
The research direction of the HIV-Specific Immunity Section of the Laboratory of Immunoregulation is focused upon on typography gill mechanisms of immunity to HIV. The laboratory is staffed by PhD and chris MD scientists with extensive experience in the humoral or cellular immune response to HIV. In addition we have close collaborations with investigators at the NIAID Vaccine Research Center in the area of structure-function relationships of HIV-specific antibodies. Our laboratory is well equipped with multi-laser flow cytometers, automated magnetic sorters, and imaging equipment to carry out this project. Our laboratory is on typography, supported by essay technology the research budget of the Division of Intramural Research. Over the past 4 years we have received supplemental funding from the Intramural AIDS Targeted Antiretroviral Program (IATAP).
This will continue for the next 2 years. We have also received funding from the Office of essay gill, AIDS Research for work on the humoral response to HIV. Positions and Employment. 1985 - 1992 Lecturer, Department of Genito-Urinary Medicine, University College London, UK 1989 - 1991 Visiting scientist, Howard Hughes Medical Institute, Columbia University, New York, NY 1992 - 1998 Staff scientist (Director of Research), Centre d’Immunologie, Marseille, France 1999 - 2003 Reader, Section of Infectious Disease, Imperial College London, London UK 2003 - Professor of immunology, Department of Pathology, University of prompts, Oxford, Oxford UK. Other Experience and Professional Memberships. 1995 - Member, American Association for the Advancement of Science 1998 - Member, American Society for Microbiology 1998 - Member, British Society for essay on typography Immunology, UK 1998 - Member, Society for General Microbiology, UK 2005 - Member of the Pasteur Institute Paris France Scientific Evaluation Committee. Chris Thesis! 2005 - Member of the Vaccine Research Center (VRC) NIH Board of Scientific Councilors 2007 - Member of BMGF Vaccine Monitoring Centre SAB 2008 - Member of NIH HIVRAD/Novartis Neutralizing Antibody Vaccine Consortium SAB 2009 - Member of study section for essay gill Swedish Program for International Development 2010 - Member of essay transport, NIH study section for the B cell Immunology Partnership Program for HIV-1 Vaccine Discovery (U19) C. Selected Peer-Reviewed Publications. Most relevant to the current application. Martin N, Welsch S, Jolly C, Briggs JA, Vaux D and Sattentau QJ. (2010) Virological synapse-mediated spread of human immunodeficiency virus type-1 between T cells is sensitive to entry inhibition.
J. Virol. 84: 3516-3527. Gonzalez N, Bermejo M, Calonge E, Jolly C, Arenzana-Seisdedos F, Pablos JL, Sattentau QJ and Alcami J. (2010) SDF-1/CXCL12 production by mature dendritic cells inhibits the propagation of X4-tropic HIV-1 isolates at essay on typography, the dendritic cell-T-cell infectious synapse. J Virol. 84:4341-51 Groot F, Welsch S and Sattentau QJ. (2008).
Efficient HIV-1 transmission from chris hoover thesis, macrophages to essay on typography gill, T cells across transient virological synapses. Blood 111: 4660-4663. Essay! Sowinski S, Jolly CJ, Berninghausen O, Purbhoo, MA, Chauveau A, Kohler K, Oddos S, Eissmann P, Brodsky FM, Hopkins C, Onfelt B, Sattentau QJ and Davis DM (2008). Membrane nanotubes physically connect T cells over long distances presenting a novel route for HIV-1 transmission. Nature Cell. Biol. 10: 211-219. Jolly C and Sattentau QJ (2007).
HIV-1 assembly, budding and essay on typography cell-cell spread in T cells takes place in oil characterization thesis, tetraspanin-enriched plasma membrane. J. Virol. 81:7873-7884 Jolly C and Sattentau QJ (2007). Adhesion molecule interactions facilitate human immunodeficiency virus type-1-induced virological synapse formation between T cells. J. Virol.
81: 13916-13921. Jolly C, Mitar I and Sattentau QJ (2007). Requirement for an intact actin and tubulin cytoskeleton for efficient HIV-1 assembly and spread. J. Virol. 81: 5547-6 Jolly C and Sattentau QJ. Human Immunodeficiency virus type-1 virological synapse formation in T cells requires lipid raft integrity.
J. Virol. 2005 79: 12088-12094. Jolly C, Kashefi K, Hollinshead M and Sattentau QJ (2004) HIV-1 cell-to-cell transfer across an Env-induced, actin-dependent synapse. Essay! J. Exp Med. 199: 283-193. Additional recent publications of chris, important to the Field (in chronological order) Wegmann F, Krashias G, Luhn K, Laamanen K, Vieira S, Jeffs SA, Shattock RJ and Sattentau QJ (2011) A novel strategy for inducing enhanced mucosal HIV-1 antibody responses in an anti-inflammatory environment.
PlosOne 6: e15861. Kong L, Sheppard N, Stewart-Jones G, Robson CL, Chen H, Xu X, Krashias G, Bonomelli C, Scanlan CN, Kwong PD, Jeffs SA, Jones IM, Sattentau QJ (2010) Expression system-dependent modulation of HIV-1 envelope glycoprotein antigenicity and immunogenicity. Gill! J. Mol. Biol. Aug 25th epub. Sports! Krashias G, Simon K, Wegmann F, Kok WL, Ho LP, Stevens D, Skehel J, Heeney JL, Moghaddam AE and Sattentau QJ. (2010). Potent adaptive immune responses induced against HIV-1 gp140 and influenza virus HA by a polyanionic carbomer. Vaccine 28: 2482-2489.
Zanetti G, Briggs JAG, Grunewald K, Sattentau QJ and Fuller SD. SIV spike glycoprotein structure in situ determined by cryo-electron tomography. Plos Pathogens August 2 (8): e83 Sheppard NC, Bates AC and Sattentau QJ (2007). A functional human IgM response to gill, HIV-1 Env after immunization with NYVAC-HIV C. AIDS 19: 524-527. [Outside research support redacted from sample.] PHS Fellowship Supplemental Form, Research Plan.
[Our Web version of best way to an opinion essay, this sample application can't reproduce the layout of the actual PDF forms. The attachments are included below. In the original version, these attachments met the on typography, page limits.] Understanding the basis of an immune response that controls infection or provides sterilizing immunity remains a major goal in the search for effective vaccines or immunotherapies for HIV. Antibodies (Abs) induced by candidate vaccines to the surface envelope glycoprotein have not neutralized a broad array of primary virus isolates. For this reason, eliciting a cytotoxic cellular response has been the primary goal in most recent vaccine trials. Best Write Essay! However, this approach has not been successful in containing viral replication in vaccinees that have become HIV-infected. Antibody-dependant cellular cytotoxicity (ADCC) has been shown to essay, mediate sterilizing immunity against challenge with pathogenic simian immunodeficiency virus [Hessel 2007]. In ADCC, Fc-bearing Abs bind viral epitopes coating an infected CD4+ target T cell and an Fc receptor bearing effector, most commonly natural killer cells (NKs), bind the Ab and use perforin to deliver granzymes which induce apoptosis in the target. We want to study ADCC in thesis, infected patients to understand the magnitude and characteristics of the best responses achieved by natural infection.
First, we will compare ADCC mediated by the sera of gill, a cohort of patients using a granzyme B cytotoxicity assay developed in our lab. Based on these findings, we will select the sera of patients with the most ADCC, generate monoclonal Abs (mAbs), and characterize the mAbs based on epitope specificity, affinity, potency, breadth, IgG isotype, and Fc type. We will also evaluate whether ADCC is disparate from classical neutralization. Finally we will use microscopy to examine the synapse between effectors, Abs, and targets. The outcome of this research will provide insight into the characteristics of Abs that mediate ADCC that are likely important goals in essay of evolution, the design of HIV vaccines or immunotherapies.
Hypothesis: Antibody-dependent cellular cytotoxicity (ADCC) is a function that has been shown to mediate protection from essay, lentiviral infection. We hypothesize that variations in ADCC activity of sera are dictated by the amount, specificity, and subclass of HIV-specific antibodies. Aim 1 : Characterize the potency of sera of chris thesis, HIV-infected individuals in on typography gill, ADCC. In ADCC, Abs bind viral epitopes that are presented by infected CD4+ T cells. NKs expressing an Fc receptor bind the Fc domain of the Ab and use perforin to essay on the, deliver granzymes to the HIV-infected cell. On Typography Gill! Subsequently, granzymes induce apoptosis within the cell. Essay! Our lab has developed a flow cytometric assay that measures granzyme B delivered to an HIV-infected CD4+ target T cell. We will classify ADCC by the percent of gill, target cells receiving granzyme and the elimination of targets as defined by residual percent of targets expressing p24, HIV capsid. Compare the serum of HIV+ individuals with various rates of progression and viral loads to theory, determine which contain Abs capable of mediating the highest levels of ADCC.
Compare the ADCC and neutralizing activity of patient sera. Aim 2 : Characterize the specificity and breadth of antibodies with ADCC activity. Our laboratory has panels of essay on typography gill, NAbs derived from patients with known serum neutralizing or ADCC-mediating activity. Determine whether recognition of specific epitopes is required for ADCC. Define the essay transport technology, breadth of the polyclonal sera by its ability to mediate ADCC in CD4+ T cells infected by different clades of HIV. Titer serum total IgG, IgG1, and IgG3 binding infected CD4+ T cells. Aim 3 : Characterize the structure and function of the target-effector synapse.
Using both fixed and live cell laser scanning confocal microscopy (LSCM), transmission electron microscopy (TEM) and cryo-electron microscopy (cryo-EM) and tomography, we will examine the synapse formed between NK and other cells with potential ADCC activity (macrophages and neutrophils) and infected target cells. We will specifically investigate: The structure of a functional ADCC synapse. The kinetics of essay on typography gill, ADCC function in real time and sports prompts its relation to antibody type and essay on typography specificity. A role for antibody-dependent cell-mediated phagocytosis (ADCP) in elimination of HIV-infected cells. d. Oil Characterization Thesis! Receptors and effector molecules central to ADCC activity against HIV infected cells. Since being identified as the causative agent of gill, AIDS in 1983, HIV infection has reached epidemic proportions with an estimated 2 million people worldwide dying from AIDS each year 1- 3. There is essay on the theory of evolution, more public awareness about how to on typography gill, prevent HIV infection but with the growing incidence of HIV infection and essay prompts the limitations of on typography gill, current antiretroviral therapy, there is an imperative need for a vaccine. One of the most critical areas of mla format, research on essay gill vaccines for HIV is the search for correlates of immunity. At present the only method for determining the potential benefits of a vaccine with regard to protection from essay of evolution, infection or lowering of viral load upon infection remains very large clinical efficacy trials.
Determining correlates of immunity would tremendously accelerate the development process by permitting some estimation of clinical efficacy prior to a Phase III trial. Reliable correlates would permit screening of vaccine candidates at a much earlier point in development with smaller numbers of on typography gill, participants. Over the past 20 years, research into correlates of for essays quoting, immunity to HIV have largely focused on CD8+ cytotoxic T lymphocytes (CTLs) or neutralizing antibodies (NAbs). In addition to neutralization, antibodies (Abs) may fight viral infections by antibody-dependent cell-mediated phagocytosis (ADCP) or by antibody-dependent cellular cytotoxicity (ADCC). In ADCC, Abs bind viral epitopes presented on infected target cells and Fc? receptor-bearing effector cells such as natural killer cells (NKs) recognize and bind the Fc? region of these Abs. Upon cross-linking of the on typography gill, Fc? receptor, NKs release perforin that punctures the target’s cell membrane and deliver proteases such as granzyme B (GrB) which induce apoptosis. ADCC has become a major focus within the field following two important studies highlighting its potential importance in way to an opinion essay, providing immunity to HIV. A recent study by Burton and colleagues demonstrated that neutralizing Abs with mutated Fc? receptor binding regions were less effective in protecting macaques from HIV whereas Abs with impaired complement binding conferred protection to macaques, suggesting that ADCC may be more important in preventing infection than neutralization or complement activation alone 4. Prior to essay, this study, it was widely accepted that infection of a single cell would lead to integration and chronic infection. Oil Characterization! However, because ADCC mediated effects are on virus infected cells and not free virions, this study provided indirect evidence and proof of the concept that ADCC can eliminate lentivirus-infected cells in vivo. In addition, some recent data in human efficacy trials has also greatly increased interest in ADCC as an important component of vaccine induced immunity to HIV.
In September 2009 data from the phase III RV144 (Thai) vaccine trial were released and showed that the prime-boost combination of ALVACR HIV and AIDSVAXR B/E reduced HIV infection. This vaccine regimen was known to not elicit a strong cellular or NAb response. It does, however, elicit binding antibodies that have suggested to essay gill, the field that the protective efficacy is likely mediated through ADCC 5. Essay! Our laboratory is part of a large international effort to characterize ADCC antibodies from this trial in order to determine whether there were higher ADCC-mediating antibodies in uninfected vaccinees compared to those that were infected. Although HIV-specific ADCC is now considered an extremely important part of vaccineinduced immunity several basic questions regarding the nature of an effective ADCC response have not been addressed. For example, the number, affinity, and specificities of antibodies necessary to mediate ADCC remains poorly understood. Our laboratory has now developed assays that can accurately measure ADCC against essay gill, HIV-infected cells. We also have assembled panels of monoclonal antibodies from oil characterization, individual patients that permit a characterization of individual antibodies that mediate HIV-specific ADCC. One of the initial goals of this project was to adapt our current killing assay to provide a more robust assay for essay gill HIV-specific ADCC. Past studies of ADCC have been hampered by the availability of chris hoover, assays that are only partially quantitative. There are four primary methods that people use to measure ADCC 6. The most prevalent is the chromium release assays (CRA), which is essay gill, highly variable, does not specifically account for the mechanism of killing, and oil characterization thesis is insufficiently quantitative 7. The ADCVI assay incubates targets, serum, and essay effectors for 24 hours and the supernatant is applied to the B95-8 cell line for 5-7 days. The endpoint is reduction of viral infection of the B95-8 cells 8. This method measures inhibition of essay on the theory of evolution, viral production and subsequent infection but its disadvantages lie in the length of the essay on typography gill, assay and failure to measure killing of infected cells.
Another assay is the intracellular cytokine staining ADCC (ICS-ADCC) assay measures IFN-?, the an opinion essay, primary cytokine responsible for the antiviral activity of many cells including NKs, CTLs, Th1, and essay dendritic cells 9. The advantages of this assay are that multiple effector functions can be measured and all cells and Abs are from the same patient, but it is still a proxy that does not measure HIV-infected cell killing. The rapid and mla format quoting fluorometric antibody-dependent cellular cytotoxicity (RFADCC) assay directly measures cell killing but requires whole protein 10. While use of protein pulsed targets is a sensitive method, it likely places artificially high amounts of protein on on typography the surface of infected cells, potentially overestimating the essay on the of evolution, efficacy of ADCC. Recently our lab has developed a highly quantitative flow cytometric assay to measure GrB cytotoxicity using primary HIV-infected CD4+ T cells as targets and on typography autologous NKs as effectors. This is an adaptation of an assay for sports essay measuring the cytotoxic activity of CTLs developed in our lab11. This assay measures the cleavage of a GrB substrate, and eliminates background cell death using a Live/Dead stain to produce accurate quantitative and highly reproducible results. In brief, CD4+ T cells are stimulated for 3 days and HIV infected. Autologous, negatively selected NKs are used as effectors. Effectors and targets are labeled with surface dyes so that they can be discriminated.
NKs, targets and serum are incubated in the presence of a granzyme cleavable fluorescent substrate. Granzyme delivery to the target cell and elimination of HIV-infected targets can then be measured by flow cytometry (Figure 1). Cellular GrB content. GrB cleaves GranToxiLux substrate (FITC), indicating the amount of GrB present within a target cell (Pacific Blue). Elimination of HIVSF162-infected cells. Live targets are stained for intracellular HIV capsid, p24 (PE). On Typography Gill! Some infected cells show downregulation of CD4 (PE-Cy7). Essay! A reduction in the number of p24 stained cells is represented as Infected Cell Elimination (ICE) and is an indication of essay, cell death. In the example above, ICEInfected+Effectors+Serum = [(58.2-25.8)/58.2] x 100 = 54.9%. There are several advantages to this assay over hoover thesis, older assays that use mismatched cell lines.
Because NKs lyse target cells in the context of MHC mismatch and essay cause nonspecific killing, autologous cells are used in thesis, our assay to reduce background. 12. In addition, because NK function is diminished in HIV-infected patients, we use the essay, NK cells and thesis CD4+ T cell targets from uninfected patients in our assay as a readout for the effectiveness of a given serum sample in mediating ADCC 13, 14. We observed variations in NKs from normal individuals in their ability to mediate ADCC and choose donors that provide optimal signal to noise ratio that we will use to compare the serum of HIV+ individuals (Figure 2). The targets used in this assay are likely closer to the in on typography, vivo situation given autologous HIV-infected targets are used in the place of antigen pulsed cell lines.
Other advantages include our assay’s ability to measure the on the, mechanism of ADCC (granzyme delivery) and essay on typography the outcome (death of sports prompts, infected cells) (Figure 2). Essay Gill! MHC-1 is down-regulated in virally infected cells, and the loss of MHC-mediated killer inhibitory signals permit NKs to kill a cell 15, 16. We control for this by measuring the granzyme content of targets that are incubated only with effectors and sera from HIV-uninfected volunteers. For a positive control we are using a novel anti-CD3 Ab that will mediate ADCC to all CD3-bearing targets (Figure 3). With this Ab we have already demonstrated (1) the sensitivity and dynamic range with which we are able to measure ADCC, (2) Abs mediate ADCC in a dose dependent manner, and (3) GrB delivery is way to write an opinion essay, proportional to cell death. Aim 1: Characterize the magnitude of ADCC in HIV infected individuals . Rational : There is considerable controversy regarding the on typography gill, magnitude of ADCC activity in sera from HIV+ individuals and how it correlates with disease progression.
It has been suggested that sera from elite controllers exhibit greater ADCC activity which correlates with CD4+ T cell count 17, 18 while others have sighted no correlation between ADCC and disease progression 19, 20.We will compare the chris hoover thesis, serum of HIV+ individuals with various rates of progression and viral loads to on typography, determine which contain Abs capable of mediating the highest levels of ADCC. We will also compare the ADCC and neutralizing activity of patient serum to evaluate whether or not ADCC and neutralization always occur together in polyclonal sera which is yet to be determined 21, 22. Methods : Over the past several years, in collaboration with the Vaccine Research Center (VRC) our laboratory has focused upon recruiting HIV-infected patients with broadly crossneutralizing antibodies. Essay Technology! We have extensive clinical data and essay on typography neutralization data for all of the essay prompts, patients. With the data we generate from our ADCC assay we are able to gill, compare multiple indicators of immunity with ADCC. Expected Outcomes/Interpretation : Based on in vitro and passive transfer macaque studies, we expect LTNPs to exhibit greater ADCC than progressors. Mla Format Quoting! Our current knowledge attributes LTNP delayed progression to superior CTL cytotoxicity but our data may show ADCC also to be a significant factor 11. Potential Complications : We do not foresee any difficulties in accomplishing this objective.
Aim 2: Characterize MAb from serum identified as having potent ADCC activity with regard to specificity. Rational/Methods : HIV-infected cells express Envelope (Env) protein on their surface which is recognized by essay on typography NAbs and presumably by ADCC-mediating Abs. We will use standard Nabs (including b12, 2F5, 2G12, 447-52D, 17B, and VRC01) and patient derived MAbs to probe specificities required for technology ADCC. On Typography! An ideal HIV vaccine would be able to protect against diverse virus strains so it is important to determine whether Abs raised against one clade of virus would be able to mediate ADCC in CD4+ T cells infected by different clades. We will use multiple strains of virus from thesis, clade A/E, B, and C to infect CD+ T cells and compare killing of on typography gill, target cells in the presence of hoover thesis, serum pooled from clade B HIV+ patients. IgG1 and IgG3 are the predominant IgG isotypes with roles in gill, ADCC. It is possible that variations in ADCC activity of sera are mediated by variations in IgG subclass. Sports Essay Prompts! We will use infected CD4+ T cells, apply sera with various levels ADCC at the same dilutions used in our killing assay, and measure total bound IgG, IgG1, and on typography gill IgG3 by flow cytometry.
We believe flow cytometry is a better way of on the, measuring titers of binding Abs as compared to ELISA because Abs will recognize all possible epitopes on the surface of an infected cell in their natural conformation. Expected Outcomes/Interpretation : Determining the association between ADCC activity and IgG subclass may provide insights into the necessity of essay on typography gill, eliciting a specific class of IgG via particular antigens and adjuvants. Potential Complications : We now have robust assays for essay on the theory subclass binding and do not expect complications with this portion of the project. Aim 3: Evaluation of the target-effector synapse. Rational : Using microscopy to study ADCC is important because it will help us define structural and kinetic parameters and will relate structure to on typography gill, function.
Multiple mechanisms for cell-to-cell spread of HIV have been described but few have described inhibition of viral transmission or targeted cell death 23. To date there has been no study using microscopy to best way to, investigate ADCC in HIV using NKs or macrophages as effectors and gill CD4+ T cells as targets. In 1994 one group designed a bispecific antibody with anti-CD4 and essay anti CD3 heavy chains that would redirect CD8+ T cells to kill HIV infected targets 24. With fluorescent microscopy they observed after 1 hour of incubation that 24% targets had formed conjugates with CD8+ T cells although many more cells had lysed. At 2 hours 50 - 80% of gill, targets were conjugated and undergoing lysis and by 24 hours all targets had been lysed. However, the bispecific antibody may not be representative of ADCC with naturally occurring HIV-specific antibodies. In addition, the structure of the NK to HIV-infected target synapse was not examined. Methods : The lab of hoover thesis, Quentin Sattentau has focused on examining the synapse of cells involved in HIV infection.
They were the first to essay, demonstrate HIV-1 spread between T cells via virological synapses and nanotube structures for viral transmission between CD4+ T cells 25. Mla Format For Essays Quoting! They have recently demonstrated that macrophages can infect CD4+ T cells every 6 hours and that cell-tocell transfer of virus is 10-fold more efficient than free virus spread 26. To study the kinetics of ADCC in real time we will use time-lapse laser scanning confocal microscopy (time-lapse LSCM). Cell populations will be negatively selected from PBMCs using magnetic cell sorting. HIV-infected CD4+ T cells will be mixed with patient serum or Abs of multiple specificities (see Aim 2) and NKs to evaluate ADCC; or neutrophils/monocytes to evaluate antibody-dependent cell-mediated phagocytosis (ADCP) and applied to poly-L-lysine coated cover slips. Cover slips will be fixed in paraformaldehyde, quenched with NH4Cl, permeabilized with Triton-X, and stained for granzyme B, IgG, CD4/CD16/CD14 with fluorescent probes. After mounting, images will be acquired every minute for 6 hours using a Zeiss Pascal Axivert 200M scope. We will use transmission electron microscopy and essay on typography gill tomography (TEM) to essay on the of evolution, generate higher resolution static images of the ADCC functional synapse. On Typography Gill! For TEM targets, serum, and effectors will be incubated for approximately 1 hour, fixed in paraformaldehyde and glutaraldehyde, and stained with osmium tetroxide. Samples will be embedded in epoxy resin, cut, and transport technology imaged using a 300kV FEI Technai TF30 27.
If necessary cryo-EM will also be used for structural studies with the essay on typography, aid of Oxford collaborator Kay Grunewald. Sports Essay! These experiments will be completed using the pathology department’s core bioimaging facility and I will take department’s Advanced Light Microscopy and Electron Microscopy course in preparation for this research. Expected Outcomes : We expect to elucidate the kinetic and functional relationship between ADCC components and answer questions such as what density of Env epitopes are needed to recruit effectors, how many effectors are recruited per target. We do not foresee any difficulties in accomplishing this objective. The results of these studies will include novel MAb with possible therapeutic value; a new tool for the study of ADCC with potential utility for HIV vaccine trials; and increased understanding of basic HIV immunology. Essay On Typography! In the wake of the RV144 trial, ADCC has become a major focus in the field of HIV-specific immunity. We are very well positioned to carry out essay prompts, these important studies because of our large collection of cells, sera, and essay gill detailed clinical data from patients that are part of the NIAID HIV clinic, and both labs’ extensive experience in this area. Samples are collected under the NIAID IRB-approved protocol 02-I-0086. Protections for human subjects are in essay prompts, accordance with NIAID Division of essay on typography, Intramural Research policies.
Phlebotomy may cause some discomfort, and occasionally some bleeding or bruising at the site. On rare occasions, people faint while having blood drawn. Blood drawing will not exceed 150 milliliters (30 teaspoonfuls) on any day or visit. Essay Prompts! In the course of on typography, any time frame during this study, the total amount of blood will not exceed 10.5mL/kg, or 550 milliliters over any 8-week period, which is within the safety guidelines for blood drawing practiced at the Clinical Center. Like standard phlebotomy, leukapheresis is occasionally associated with local bleeding and bruising at the site of needle insertion.
Lightheadedness can also occur, and people faint on rare occasions. Occasional tingling in the arms and legs can occur with the technology, procedure and can usually be relieved by chewing a calcium-rich tablet that the staff will provide. In addition, on very rare occasions (less than 1 in on typography gill, every 1,000 times), malfunction of the leukapheresis machine can result in blood loss of up to 450 milliliters. Trained medical personnel who are equipped to deal with any of these potential problems will always be on hand during the procedure. Inclusion of Women and chris Minorities. Gender, Ethnicity and Race Consideration. This study involves techniques and essay on typography gill procedures that are currently approved for various indications. Male and female adult patients (18 years of age or older) will be enrolled without preference to gender, race or ethnicity from the pool of patients that receive care in essay technology, Clinics 8 at the NIH Clinical Center. Patients will be recruited from outside the NIH community by essay advertisements posted in the Clinical Center, the NIAID web site, and advertised in other media. We will attempt to facilitate minority recruitment using existing NIAID community outreach efforts involving local clinics in the Washington, DC metropolitan area.
In adult HIV-infected patients planning to interrupt HAART, patients will be fully counseled prior to entry into oil characterization thesis, the study as to on typography, the potential risks of sports, discontinuing HAART therapy and the potential benefits of continuing HAART therapy, in gill, the setting of a documented virologic and/or immunologic response. Patients who do not fully comprehend these potential risks of HAART cessation, in the opinion of the chris hoover, study team, will not be offered participation in the study. Patients stopping HAART therapy will be monitored closely for virologic and immunologic relapse and will be offered therapy, based upon the algorithm for on typography safety outlined above. Target enrollment for this study is 300 individuals and is currently ongoing. Inclusion of essay, Children (and Pregnant Women) Inclusion of Children and essay on typography gill Pregnant Women. This study will be confined to HIV-infected adults greater than or equal to 18 years of age. As an observational study there is sports prompts, no therapeutic benefit or clinical intervention resulting from participation in this trial, thereby children are not being denied the potential benefits of essay on typography gill, a research intervention. We also do not have the historical research database and in vitro findings on HLA linkages with LTNP in the pediatric population to make valid comparisons within that cohort.
The potential risks of oil characterization thesis, withdrawing therapy in minors have not been studied and therefore may present unknown additional risks to essay on typography gill, that study population. Best Write Essay! Comparable units specializing in pediatric HIV are available at the NIH to address this topic, should preliminary results from this study warrant further evaluation in the pediatric population. The safety of discontinuing antiretroviral therapy has not been established during pregnancy and nursing, and current guidelines continue to advise HIV-positive women to on typography gill, take antiretroviral therapy to essay transport technology, prevent viral transmission to the child during that time period. Therefore, pregnant or nursing women will be excluded from on typography gill, this study. Any woman of childbearing potential must have a negative pregnancy test within two weeks prior to study entry. Essay Prompts! All participants who engage in essay on typography gill, heterosexual intercourse must use two forms of reliable birth control for the duration of the study. When I entered the essay, NIH/Oxford/Cambridge Scholars program I identified Mark Connors and Quentin Sattentau as two HIV researchers with whom I would like to train with because of their focus on gill HIV immunology, vaccine design, and their expertise in basic immunology techniques, flow cytometry, and microscopy. I initiated contact between the two individuals and they agreed to start a collaborative project with me. After studying NAbs with Mark Connors for one summer I decided I wanted to focus on on the theory antibodies that mediate ADCC. On Typography Gill! I outlined the facets of ADCC I wanted to investigate then received feedback from both mentors. I subsequently wrote the essay prompts, proposal that was edited by both mentors and Mark Connors made final revisions.
I meet multiple times during the on typography, week with Dr. Connors to discuss my research progress and to oil characterization, plan future experiments. We also conduct quarterly phone conferences with Dr. On Typography Gill! Sattentau, maintain a regular email correspondence, and meet at theory of evolution, conferences. Communication will continue in the same manner while I am in Quentin Sattentau’s lab. Selection of Sponsors and Institution. Selection of Institution. I decided to join Columbia’s MSTP program because of on typography, its tradition of thesis, exemplary medical education, medical research, and on typography unparalleled support offered to students from program administrators. Oil Characterization Thesis! Because of my previous research experience with HIV I knew I wanted to continue research in essay gill, this field.
I decided to chris thesis, join the NIH/Oxford/ Cambridge Scholars Program to complete my dissertation research because it would provide me access to a more extensive network of scientists than Columbia could. Oxford has a particularly strong network of HIV investigators and the structure of its graduate program allows for a particularly specialized education. As a graduate student in essay gill, the pathology department I am allowed to select seminars and accelerated 1-day to 1-week courses that allow me to attain skills quickly and immediately implement them into my research. In addition, the NIH/Oxford/Cambridge Scholars program provides funding for students to take courses offered by any institution giving me access to virtually any educational opportunity necessary. In addition, through NIH’s Foundation for Advanced Education in the Sciences I’ve had the opportunity to transport technology, take courses that will help me towards my goal as in infectious disease investigator such as the Vaccine Development course I completed. The NIH/Oxford/Cambridge Scholars Program is designed so that students work with a mentor at gill, NIH and one at a university. When I joined the program I searched for HIV investigators throughout NIH and Oxford and identified Mark Connors and Quentin Sattentau as potential mentors because I wanted to conduct research that had direct implications in an opinion essay, vaccine development.
Quentin Sattentau was studying how to elicit effective anti-HIV neutralizing Abs (NAbs) for vaccines and adjuvants to stimulate systemic and mucosal immunity. Through my search at essay gill, the NIH I identified Mark Connors as a potential collaborator though he and Quentin had never collaborated before. I believed Dr. Connors’s expertise in NAbs of long-term nonprogressors (LTNPs) would complement Dr. Sattentau’s vaccine development goals. In addition, Dr. Connors’s unique cohort of LTNPs would provide ready access to samples to study many facets of mla format for essays, HIV infection. Finally, I believe Dr. Sattentau’s approach to research as a PhD and Dr.
Connors’ as an MD will teach me to on typography, think about science in oil characterization thesis, a more practical way that is suited to me as a future MD/PhD researcher. I will follow the principles on responsible conduct of research as outlined in NOT-OD-10- 019. As an undergraduate at essay gill, UMBC I took a semester-long course entitled “Ethics/Integrity in Scientific Research” which first help me to understand the responsibility scientists have to one another and medical community honestly share information. As a Columbia medical student, I have taken the Hippocratic Oath to guide my patient care and I am expected to adhere to the code of ethics as outlined in thesis, student handbook. Upon joining the NIH I took an online ethics course and essay will continue to sports prompts, take refresher courses annually. At Oxford I will continue my ethics training by taking the required “Ethics: Introduction to essay on typography gill, Research Ethics”.
Since I will have access to thesis, sensitive patient data it is essential that I maintain the on typography, confidentiality of mla format, these individuals. When I started research at NIH I participated in essay, a patient confidentiality seminar. Oxford offers an “Ethics: Confidentiality” course in best write essay, which I will enroll to better my further my understanding and ability to on typography gill, maintain ethical practices with regards to research. As a physician, I hope to alleviate an individual’s immediate suffering; and as a scientist I hope to contribute to long-term solutions that can prevent suffering. To work most effectively at the interface of medicine and science, I have endeavored to earn an chris thesis, MD/PhD.
I believe that the combined degree’s greatest value lies in the perspective that it affords: to essay gill, approach patient care with a researcher’s eye, and to sports, approach research with a physician’s focus on improving the essay, quality of life. A few years ago I conducted research on telomerase, an essay transport, enzyme that maintains healthy chromosome length. When telomerase is upregulated, as it is in essay on typography gill, 90% of carcinomas, cells can propagate indefinitely. When telomerase levels are halved, a bone-marrow failure disease called Dyskeratosis Congenita (DC) results. While looking at yeast cells with low levels of telomerase and measuring their growth rates, I noticed two things. The first was that cells with low levels of telomerase grew slowly. Secondly, these cells had abnormal morphologies, as do tumor cells. These observations led me to wonder whether the progression of transport technology, cancer in individuals with DC was slower than in essay on typography gill, patients without it because DC patients have less active telomerase.
Upon discussing this with my P.I., he acknowledged having never considered his research from that aspect. A literature search demonstrated a lack of prompts, any substantial research concerning this question. Research in biochemistry affords many new advances in medicine, but through that experience the importance of keeping a broader medical picture in essay, mind became clear to me. My goals as a physician scientist are to treat patients and conduct research that will allow me to impact patients I will never meet through therapeutic discovery; specifically serving as a creative link between science and medicine in the field of infectious disease. The training I will receive in Oxford’s doctorate program will prepare me to sports essay prompts, be a competitive investigator through a tailored curriculum, expert dual-mentoring, learning to essay, work in a collaborative environment, and by giving me extensive writing experience via paper submissions and even through this grant. These are all specific skills that would be harder and take longer for me to garner through a traditional medical student experience. The advantage I have in understanding that I want to be a physician-scientist has given me the best way to essay, opportunity to carefully cultivate these skills early in my career. Activities Planned Under This Award.
Research 80%, Seminars/Courses 10%, Presentation/Career Development/Clinical 10% Research Skills : While working in Mark Connors’s lab I have acquired specific skills such to creatively study immunology such as learning to use flow cytometry. I discuss my research progress frequently during the week with Dr. Connors. We also have phone conferences and meet with Dr.
Sattentau at numerous conferences throughout the year. Writing Skills : When I entered the program I took a grant-writing workshop sponsored by the NIH Graduate Partnership Program. I also completed a 3-day grant-writing course during my senior year of college. I will improve my writing skills through preparation of essay on typography gill, manuscripts based on my research to be submitted for publication, applying for chris hoover training grants, and submission of a report for essay on typography my transfer viva (qualifying exam). I will also complete the Scientific Writing from sports essay prompts, a Reader’s Perspective seminar. Discipline specific learning : I completed the 3-day “Vaccines: Development and Evaluation of Efficacy” course offered by FAES in February 2010. I also attended “Psychiatry and essay gill HIV,” a 2-day workshop offered by the American Psychiatric Association so I could learn to effectively interact with HIV+ patients I encounter.
I attend weekly seminar series hosted at the NIH such as the Wednesday Afternoon Lecture Series, Immunology Interest Group seminar series, and weekly lab meetings. Bench to bedside clinical training : I will attend the Clinical Research Training Program (CRTP) weekly rounds. I will also have the opportunity to be mentored by an infectious disease clinician. Presentations : I will continue to annually attend and present at pertinent Keystone Symposia and the Annual AIDS Vaccine conference. I have also had opportunities to present posters at the NIH/Oxford Cambridge Scholars Program Colloquium and the Oxford Pathology Graduate Student Symposium. I have given a talk to my graduate program journal club and will have more opportunities to present my research in lab meetings. Career Development : The NIH/Oxford/Cambridge Scholars programs hosts a seminar series where leaders in transport technology, government/academic research, the pharmaceutical industry, and essay gill finance speak with us about their career. I also attended NIAID Fellows retreat that focused on career development.
Teaching : As I prepare to quoting, leave the essay, lab I will be responsible for training a new research fellow. Year 1 (continued) and Year 2: Quentin Sattentau Lab (Oxford) Research 75%, Seminars/Courses 10%, Presentation/Career Development/Clinical 10%, Teaching 5% Research Skills : While working in Quentin Sattentau’s lab I will learn to apply microscopic imaging to studying HIV infection. I will have weekly meetings to discuss my research progress. We will have phone conferences and meet with Dr. Connors at mla format for essays, various conferences throughout the year. I have also been assigned a graduate advisor within the essay on typography, pathology department but not in my specific lab who will help me to objectively evaluate my research. Writing Skills : I will improve my writing skills through preparation of manuscripts based on my research to for essays quoting, be submitted for publication and on typography gill writing reviews. I will complete the following 1-day courses offered at Oxford: Writing Skills – Paper and mla format for essays quoting Thesis; Doctoral Thesis Writing,
Discipline specific learning : I will participate in courses and Skills Workshops offered by essay Oxford including “Advanced Light Microscopy,” “Electron Microscopy,” and “Introduction to Statistics.” Bench to bedside clinical training : I will attend the Nuffield Department of Clinical Medicine Grand Rounds and chris hoover shadow an infectious disease clinician. Career Development : See above. I will also attend the 4-day Oxford GRADschool, a personal development course. Teaching : I will seek to be an undergraduate lab instructor and take the Oxford course “Teaching and Learning Skills” to prepare myself.
Year 3 and Year 4: Medical School (Columbia) In the summer after 11th grade I participated in the intensive 6-week Gene Search Program at the Catholic University of America that introduced me to molecular biology and essay on typography gill the exciting race to sequence the human genome. That summer, I learned much more than how to best write an opinion, run a gel; I learned to read and comprehend primary research articles and how to design experiments. Essay On Typography! Because of my accomplishments that summer, I was invited to participate in an extension of the Gene Search Program during my 12th grade year. This program inspired me to pursue a career in biomedical research because I began to understand how the principles I was learning in essay prompts, school fostered scientific discovery. As a high school senior, I worked with Dr. William Winter at Howard University School of on typography gill, Medicine to investigate pathways that regulate 2,3-diphosphoglycerate (2,3-DPG) levels in essay transport, red blood cells (RBC). 2,3-DPG lowers the affinity of hemoglobin for essay on typography oxygen, thereby facilitating oxygen release to tissues. Patients suffering with sickle cell anemia have high levels of 2,3- DPG.
While this effect may seem advantageous to patients with sickle cell anemia, deoxygenating hemoglobin ultimately causes red blood cells to sickle by promoting hemoglobin polymerization. Previous studies indicated that adenosine might regulate 2,3-DPG levels through an interaction with the write essay, A3 receptor. To test this idea, we indirectly measured 2,3-DPG levels by using spectrophotometry to measure a by-product of the 2,3-DPG pathway. An exciting therapeutic prospect of our research is that an antagonist of the A3 receptor may prevent the on typography, accumulation of 2,3-DPG and RBC sickling in for essays, patients with sickle cell anemia. I documented this work in a five-chapter thesis, in accordance with my high school’s Science and Technology Program, and presented it at my school’s science fair, where I was awarded third place in biochemistry. During college I spent four years working with Dr. Michael Summers to investigate Moloney Murine Leukemia Virus (MuLV) dimerization. Understanding dimerization is essay, key to mla format quoting, developing gene and antiviral therapies.
MuLV is gill, similar to HIV in that it is a retrovirus that packages two copies of its RNA genome. Packaging of dimerized RNA is mediated by interactions between the nucleocapsid (NC) domain of the assembling Gag polyproteins and a specific domain of the RNA called the Psi-site. Previous work in the Summers lab suggested a riboswitch mechanism for MLV genome packaging, where, in thesis, monomeric RNA, Psi-site UCUG segments capable of binding NC are sequestered by base pairing, but upon dimerization, become exposed to bind NC. My project was to investigate whether similar sequences in the Psi-site could also bind NC. Using isothermal titration calorimetry and nuclear magnetic resonance, we identified short RNAs (ACAG, UUUG, and UCCG) that bind NC with nanomolar affinity. Our findings suggest that: 1) binding depends on an unpaired guanosine, 2) binding is enhanced in short RNAs containing terminal phosphates, and 3) binding affinity varies by more than one order of magnitude depending on which three nucleotides lie upstream of the guanosine. The paper I co-authored (Dey, A., et. al., 2005) extends an established model for essay on typography gill genome recognition, in which the essay transport, NC domains of assembling Gag molecules interact with multiple X(i-3)-X(i-2)-X(i-1)-G(i) elements (X is a variable nucleotide) that appear to be preferentially exposed in the dimeric RNA. Essay On Typography! I presented this research at the 2004 ABRCMS and at multiple UMBC symposiums. I continued to investigate potential dimerization and NC-binding regions of the Psi-site using native and mutant MuLV RNA segments in gel-shift assays.
As expected from our riboswitch hypothesis, we observed NC binding preferentially to RNA dimers which was also confirmed by equilibrium dialysis. Hoover Thesis! Most recently our lab published the NMR structure of this dimer (Miyazaki, Y., et al., 2010). During the summer of 2005 I participated in the Weill Cornell/Rockefeller/Sloan Kettering Gateways to the Laboratory Program. On Typography Gill! I worked with Dr. Mla Format For Essays Quoting! Nikola Pavletich toward solving the crystal structure of the Saccharomyces cerevisiae Rad4-Rad23 complex bound to on typography, a DNA substrate. Rad4-Rad23 is chris hoover, a homolog of human XPC-HR23B, a DNA damage-repair protein. In its mutated form, XPC-HR23B causes the disease Xeroderma Pigmentosum. On Typography! Patients are extremely sensitive to on the of evolution, UV-light and develop skin cancer during childhood.
I identified DNA substrates to essay, be used in crystallization trials by HPLC-purifying DNA, performing gel-shift assays, and setting up crystal trays. I presented this work in a talk before the Tri-Institutional community and during posters sessions at the 2005 Leadership Alliance National Conference and at the 2005 SACNAS National Conference. In 2006 I studied telomerase assembly in the lab of Dr. Thomas Cech at the University of Colorado at Boulder. The central role telomerase plays in protecting chromosome integrity places telomerase at the heart of two common human ailments—cancer and sports prompts aging. To study how and where in the cell telomerase is essay gill, assembled from its RNA and protein components, we used the budding yeast Saccharomyces cerevisiae as a model organism. We sought to understand whether the noncoding telomerase RNA (TLC1) goes through a cytoplasmic phase during its biogenesis. One model is that a polyadenylated precursor form of TLC1 is exported to the cytoplasm where it associates with Sm proteins.
Mtr10p then plays a role in importing the TLC1-Sm complex back into the nucleus, where the mla format quoting, telomerase holoenzyme is assembled. Using cellular fractionation and real-time RT-PCR, we attempted to localize TLC1 in its various stages. During the summer between my first and gill second year of medical school I joined the lab of Mark Connors where I screened patient sera for essay transport breadth and essay on typography potency of neutralizing Abs against HIV using a TZMBL neutralization assay. We demonstrated that that progressor serum has greater breadth in potency in neutralizing HIV than Long-Term Nonprogressors (Doria- Rose, N., et al., 2009). Studying NAbs peaked my interested in other anti-viral mechanisms mediated Abs and lead to my proposed project of technology, investigating antibody-dependent cellular cytotoxicity. Each research experience has shown me the essay gill, many nuances of a research career. I learned about: 1) the importance of basic research in developing medicines, 2) the time, organization, and ingenuity it takes to conduct research, and 3) the an opinion, vital role that a physician-scientist plays in translating basic research into medical treatment. Although I’ve experienced frustrations and on typography disappointment in research, the theory of evolution, thrill of observing something new, finishing a project, or publishing a paper always outweighs these challenges.
Each successive experience only increased my eagerness to improve health care through innovative research. [Redacted from sample.] QUENTIN SATTENTAU MENTOR STATEMENT. [Redacted from sample.] [Our Web version of this sample application can't reproduce the layout of the actual summary statement. On Typography! See below for the data fields and critiques.] Center for Scientific Review Special Emphasis Panel.
Fellowships: AIDS Predoctoral and Postdoctoral. RESUME AND SUMMARY OF DISCUSSION: In this application, the oil characterization thesis, fellowship candidate proposes to investigate antibody-dependent cellular cytotoxicity (ADCC) in HIV-infected patients do determine the extent of protection that ADCC provides. The reviewers noted that the candidate is outstanding, with a strong record of research training and accomplishment. The candidate appears to be highly motivated. Her sponsors, at both institutions, are well qualified to provide the necessary training and essay the letters of oil characterization thesis, recommendation were uniformly laudatory.
Minor weaknesses of the application included the ambitious research plan to on typography, be accomplished in two years, the risk of conducting the research at essay, two sites, and the failure to address pitfalls and essay on typography alternatives of the best an opinion, research project. These weaknesses, however, did not detract from the reviewers’ enthusiasm. This training project will have a high impact in ensuring that the candidate maintains her potential to become an independent clinical researcher. DESCRIPTION (provided by gill applicant): Understanding the basis of an way to write, immune response that controls infection or provides sterilizing immunity remains a major goal in the search for effective vaccines or immunotherapies for HIV.
Antibodies (Abs) induced by essay candidate vaccines to of evolution, the surface envelope glycoprotein have not neutralized a broad array of on typography, primary virus isolates. For this reason, eliciting a cytotoxic cellular response has been the primary goal in most recent vaccine trials. Chris Hoover! However, this approach has not been successful in essay on typography gill, containing viral replication in vaccinees that have become HIV-infected. Antibody-dependant cellular cytotoxicity (ADCC) has been shown to mediate sterilizing immunity against challenge with pathogenic simian immunodeficiency virus [Hessel 2007]. In ADCC, Fc-bearing Abs bind viral epitopes coating an infected CD4+ target T cell and an Fc receptor bearing effector, most commonly natural killer cells (NKs), bind the Ab and use perforin to deliver granzymes which induce apoptosis in the target.
We want to study ADCC in infected patients to understand the magnitude and mla format for essays quoting characteristics of the best responses achieved by natural infection. First, we will compare ADCC mediated by the sera of a cohort of patients using a granzyme B cytotoxicity assay developed in our lab. Based on on typography these findings, we will select the sera of patients with the most ADCC, generate monoclonal Abs (mAbs), and characterize the on the, mAbs based on gill epitope specificity, affinity, potency, breadth, IgG isotype, and Fc type. We will also evaluate whether ADCC is disparate from classical neutralization. Finally, we will use microscopy to examine the synapse between effectors, Abs, and essay theory targets. Gill! The outcome of this research will provide insight into the characteristics of Abs that mediate ADCC that are likely important goals in the design of essay on the of evolution, HIV vaccines or immunotherapies.
Hypothesis: Antibody-dependent cellular cytotoxicity (ADCC) is a function that has been shown to mediate protection from lentiviral infection. We hypothesize that variations in ADCC activity of sera are dictated by the amount, specificity, and on typography gill subclass of essay technology, HIV-specific antibodies. Aim 1: Characterize the potency of gill, sera of HIV-infected individuals in ADCC. Aim 2: Characterize the specificity and breadth of oil characterization thesis, antibodies with ADCC activity. Aim 3: Characterize the structure and function of the target-effector synapse using both fixed and live cell laser scanning confocal microscopy (LSCM), transmission electron microscopy (TEM) and cryo-electron microscopy (cryo-EM) and tomography. Understanding the role of antibody-dependent cellular cytotoxicity (ADCC) in HIV could provide important insights for induction of this activity through vaccination. Essay! This project seeks to sports, characterize the Abs that mediate ADCC and image the essay on typography gill, functional synapse formed by write an opinion cellular components involved in ADCC with the goal of defining new goals for the development of HIV vaccines and gill therapeutics. Fellowship Applicant: 1.
Sponsors, Collaborators, and hoover Consultants: 1. Research Training Plan: 2. Training Potential: 1. Institutional Environment Commitment to Training: 2. Overall Impact/Merit : This proposal is from an applicant who is on typography gill, currently enrolled in the NIH M.D./Ph.D. Partnership Training Program and is a participant in the NIH/Oxford/Cambridge Scholars Program, which promotes joint research in an NIH laboratory and in a university research lab. Ultimately, she will receive her Ph.D. from Oxford University and complete her medical training at Columbia University. The applicant is best, highly recommended by previous research mentors and has an outstanding academic record. Her research experience, which includes four years of gill, undergraduate research, is matched by best way to write an opinion essay her productivity; she is a contributing author on two published papers and has presented her work at essay, numerous scientific venues. The research plan, which derives directly from the expertise and interests of the mentor, focuses on relationships between antibody-dependent cellular cytotoxicity (ADCC) in HIV-1-infected patients, HIV-1-associated disease progression, and antibody subtype. Additional experiments, which will draw on essay prompts the expertise of the co-sponsor, will investigate the target-effector synapse during ADCC.
These studies will be significant for their basic science and translational value. The training plan is unique in that training and research will be accomplished in essay on typography, two separate laboratories of the sponsor and co-sponsor. This proposal outlines an excellent training opportunity that will greatly benefit the applicant and her potential for establishing an independent research career. The applicant, who is currently enrolled in an M.D./Ph.D. training program, is described in three strong letters of recommendation as enthusiastic, articulate, knowledgeable, highly focused, and very inquisitive. Her undergraduate research mentor characterized her as “easily among the top two or three of the more than 200 current and essay transport former undergraduates” that he has mentored. The applicant has four years of undergraduate research experience, which involved work with the on typography gill, Moloney Murine Leukemia Virus, and research performed during a summer program at the Howard Hughes Medical Institute focused on studies of telomerase.
Technical approaches for which she was trained included cell growth assays, transformation, RT-PCR, isothermal titration calorimetry, NMR, and cryo-electron tomography. Through her prior research experience, the applicant has been included as a contributing author on two published papers and has presented eight posters and one oral presentation at various local, national, and international meetings. The applicant has an sports essay, outstanding academic record (3.97 GPA) and numerous awards and recognitions reflecting superior academic performance. The collaborative project outlined in the proposal was an essay, outgrowth of a dialog initiated by the applicant between the sponsor and co-sponsor. The applicant has provided thoughtful goals for developing a career as both a scientist and physician. 2. Way To Write An Opinion Essay! Sponsors, Collaborators, and Consultants: The applicant’s mentor and sponsor, who is currently Chief of the essay, HIV-Specific Immunity Section in the Laboratory of Immunoregulation, NIAID/NIH, is an established and productive senior investigator with research support provided by best way to the NIH. The co-sponsor is also a productive researcher with a consistent record of gill, support for essay on the theory of evolution studies related to HIV/AIDS. The research interests and expertise of the sponsor and co-sponsor will provide excellent support for the applicant’s research and training.
The expertise of the co-sponsor will be particularly relevant to studies outlined in Aim 3. The sponsor has trained an average of essay on typography, four students per oil characterization thesis, year over 20 years, suggesting an excellent training environment for the applicant. Similarly, the co-sponsor has trained nine Ph.D. trainees since 2003. During her predoctoral fellowship, the essay gill, applicant will be interacting with four post-baccalaureate students and one M.D. in the sponsor’s lab, and four Ph.D. trainees in the co-sponsor’s laboratory. The latter trainees will overlap with the applicant for one year or more. These interactions will enrich the applicant’s training. The research plan is built on studies of antibody-dependent cellular cytotoxicity in HIV-1-infected individuals. Experiments will examine the potency of transport technology, sera of HIV-1-infected individuals in ADCC (Aim 1), the specificity and breadth of antibodies with ADCC activity (Aim 2), and the structure/function of the target-effector synapse that mediates ADCC.
The central feature of the on typography, proposed research is a novel and essay of evolution quantitative ADCC assay (developed in on typography gill, the sponsor’s lab) that offers significant improvements over other ADCC assays. Prompts! Results generated from experiments in Aims 1 and 2 will have both basic science and translational value, while results from Aim 3 experiments will provide basic information about the mechanisms of ADCC. On Typography! The applicant and sponsors are well positioned to perform these studies, since the patient populations and essay theory of evolution study procedures are already in place. The applicant is participating in the NIH/Oxford/Cambridge Scholars Program, which promotes research with a mentor at the NIH and on typography another mentor at a university. Hoover! Consistent with the essay, Scholars Program, the oil characterization thesis, research training plan includes studies and training in year 1 in the sponsor’s lab at the NIH, continued studies and training in year 2 in the co-sponsor’s lab at Oxford, and essay gill years 3 and 4 in medical school. Training in each laboratory will include the essay on the theory, development of specific laboratory skills, writing skill development, discipline-specific education, bench-to-bedside clinical training, career development, and opportunities for presentation of essay on typography gill, results and teaching. Training at the NIH (sponsor) will be a continuation of the current laboratory work, with increased emphasis on independent research and critical thinking. Training at Oxford (co-sponsor) will include skills development in experiments involving infectious HIV-1, microscopic imaging, and biochemical and serological techniques (such as ELISA, gel electrophoresis, and mass spectrometry). Although the targeted patient enrollment is given as 300 HIV-1-infected individuals, the experimental plan does not provide details regarding the scope of oil characterization, studies in on typography, Aim 1. Correlates of ADCC to best essay, be studied in Aim 1 (“rates of progression and viral loads”) were not described in sufficient detail. How will rates of progression be measured? Will viral load be documented on a continuous scale or will patient viral loads be stratified?
An explanation of how the results of Aim 1 experiments will be used to guide the selection of pooled patient sera in Aim 2 would have been informative. A research project spanning the laboratories of two different principal investigators will offer diversity in training and acquired expertise, and enhance the capability of the essay on typography gill, applicant to develop an independent and productive research career. 5. Institutional Environment Commitment to Training: The resources and facilities at essay, both institutions will provide outstanding environments for training and research. A more detailed plan regarding the coordination of training plans between the sponsor and co-sponsor would be valuable, considering the distance and time difference between the two performance sites. Training in the Responsible Conduct of Research:
Comments on Format (Required): The description of RCR training is brief and does not provide a detailed plan for formal training. However, enrollment in essay on typography, two formal courses at best way to an opinion essay, Oxford (both focused on essay gill ethics) is mentioned. Comments on Subject Matter (Required): Training courses and certifications at the NIH and Oxford should provide information on way to relevant subjects. Comments on Faculty Participation (Required): Faculty participation could not be evaluated from the description of RCR training. Comments on on typography gill Duration (Required): The duration appears to sports, be adequate, although exact contact hours were not provided.
Comments on essay on typography Frequency (Required): The frequency of training could not be discerned from the description of RCR training. Fellowship Applicant: 2. Sponsors, Collaborators, and Consultants: 1. Research Training Plan: 2. Training Potential: 2. Institutional Environment Commitment to best an opinion, Training: 1.
Overall Impact/Merit : This is an outstanding proposal from a highly qualified applicant. The applicant has an exemplary scholastic record, strong research history and outstanding letters of reference although she lacks a first author publication. The applicant has chosen two superb sponsors with complementary skill sets. In fact, the applicant brought these two investigators together herself. Both sponsors have excellent training track records and both are actively involved in the HIV arena. The primary goal of this project is to characterize antibodies that mediate antibody-dependant cellular cytotoxicity (ADCC). The Connors lab at the NIH has developed a novel flow cytometric-based assay to measure granzyme B delivered to essay, an HIV infected cells. Using this novel assay the applicant plans to thesis, compare ADCC in essay, HIV+ subjects with various rates of way to essay, disease progression and to define the specificity and breadth of antibodies with ADCC activity (Aim 1 and essay on typography gill 2).
The third Aim to characterize the structure and essay prompts function of the target-effector synapse will be carried out at Oxford under the guidance of essay, Dr. Sattentau. This aim will use fixed and essay prompts live cell laser scanning confocal microscopy and TEM to examine the gill, samples between NK and other cells with ADCC activity. If the applicant is successful the data from essay prompts, this project will be highly important for HIV vaccine development. The research project however is essay on typography gill, quite ambitious and no timeline is given. Sports Essay! It appears that the applicant plan to spend 1 more year in the Dr. On Typography! Connors lab and 1 year in Dr. Best Way To Write An Opinion! Sattentau lab making it a very busy two years if the applicant plans to finish all the work they outlined. Despite the on typography gill, highly ambitious nature of the proposed research project the overall project goals warrant further study and the applicant herself is of the highest caliber making this application worthy of sports essay prompts, funding. The applicant has a very strong research background dating back to high school where she participated in on typography, the Gene Search Project at Howard University Hospital.
As an undergraduate at UMBC she was a member of the Meyerhoff Scholars Program funded by the HHMI. The applicant has coauthored 2 papers and is the first author on 8 posters and 1 oral presentation. The applicant has extremely strong letters of recommendations, one of which is from a Nobel Prize winner. Exemplary scholastic record and many community-based volunteer positions. 2. Sponsors, Collaborators, and Consultants:
Two suburb sponsors are listed. Best Write An Opinion Essay! Dr. Connors, Chief, HIV-specific Immunity Section NIAID, has a long track record of high quality research and publications and a solid list of successful trainees (4/year for 20 years). The other sponsor, Dr. Sattentau, a Professor of Immunology and Pathology at essay, the University of Oxford, is equally meritorious with over 151 published manuscripts and solid funding record. Dr. Sattentau, also has a established track record of training PhD student (n=9) and post-docs Both sponsors have well established complementary HIV research programs that will support the applicants proposed research project. The planned research, if successful, could provide important information about of the role of ADCC in HIV infection. Essay Theory Of Evolution! The project is well outlined and there are sufficient preliminary data to support the feasibility of the project.
The plan is to utilize a novel recently developed (Connors lab) quantitative flow cytometric assay to measure GrB cytotoxicity and essay on typography determine which Abs are capable of essay transport technology, mediating the highest level of ADCC. Using this technique the applicant plans to characterize the potency of sera from subjects with different rates of disease progression, characterize the on typography gill, specificity and best write an opinion essay breadth of antibodies with ADCC activity. Using advance microcopy techniques (fixed and live cell laser scanning confocal microcopy and electron microscopy) the applicant plans to examine the essay on typography gill, structure and function of the target-effector synapse. This work will be done under the guidance of mla format quoting, Dr. Sattentau who is an essay, expert in studying the chris, virologic synapses. This is essay gill, a highly ambitious grant.
It may be difficult for best way to the applicant to finish all the gill, proposed work especially the work proposed in Aim 3. The applicant is calling for 300 subjects but no power calculation is thesis, shown to support that many subjects. A timeline for essay gill the proposed work would be helpful. The applicant does not anticipate any issues nor are any alternative approaches outlined. Chris! This maybe a little naive. The research training plan is essay gill, well laid out by mla format for essays quoting year. The plan includes research, writing skill (grant writing course), presentation, career development, teaching (both in class room training of gill, new personnel) and discipline specific learning (i.e. in house courses and seminar series, lab meetings). Letters describing each institution’s role in training the applicant were provided. Applicant states that she will interact on daily basis with section head and other senior laboratory member on a daily basis. Students also make formal presentations and attend weekly LIR, meeting, NIAID grand rounds, clinical center Grand Rounds and HIV journal club. The training plan indicates that the applicant will be at the NIH for one year and best way to write an opinion Oxford for one year and then back to on typography gill, medical school for the last 2 years.
This raises some concerns regarding the oil characterization, duration of the funding (3 vs. 2 years). 5. Institutional Environment Commitment to Training: The work will be conducted at the NIH/NIAID and University of Oxford. Both institutions are of the on typography, highest caliber with access to sports essay prompts, all the required resources. Oxford has a program in place for graduate students to essay gill, take accelerated courses and the NIH Foundation for Advanced Education in the Sciences also has specialized courses that will facility the applicants goal to become an infectious disease investigator. Training in chris hoover thesis, the Responsible Conduct of Research: Comments on Format (Required): On line ethics course at the NIH and essay formal classroom course at Oxford. Comments on essay on the theory of evolution Subject Matter (Required):
Applicant states they will follow principle of research outline in NOT-OD-1-019. The applicant has taken semester long course on Ethics/Integrity in Scientific research. The applicant took another Ethic course upon entering the NIH. Oxford offers a similar course. Comments on Faculty Participation (Required): No comment on on typography gill faculty participation. Comments on Duration (Required): Comments on Frequency (Required):
Online course and annual refresher at the NIH. Additional Comments to oil characterization thesis, Applicant (Optional): Fellowship Applicant: 1. Sponsors, Collaborators, and essay on typography gill Consultants: 1. Research Training Plan: 2. Training Potential: 1. Institutional Environment Commitment to Training: 1. Overall Impact/Merit : This proposal comes from an excellent candidate who is part of the NIH/Oxford/Cambridge Scholars program, carrying out her PhD research under the guidance of transport technology, two excellent researchers and mentors, Drs.
Connors (NIH) and Satentau (Oxford) and will complete her MD studies at Columbia. The primary hypothesis of essay on typography gill, this proposal is that ADCC activity levels in sera are dictated by the amount, specificity, and subclass of HIV-specific Abs. Transport Technology! This represents a highly relevant area of essay, research that will be tackled through 3 straightforward and well-planned aims – (i) characterize the potency of sera from individuals at different stages of HIV infection in ADCC (comparing to clinical status and chris hoover thesis Ab neutralization), (ii) characterize the specificity and on typography gill breadth of transport, Abs mediating ADCC (comparing the essay gill, sera to panels of for essays, known neutralizing Abs), and (iii) characterize the essay on typography gill, ADCC synapse. This will involve state-of-art approaches established in way to write an opinion essay, the sponsors’ labs, allowing the applicant an excellent chance to essay gill, perform this research. It is a clear and well-written proposal detailing a strategy to carry out relevant research in an excellent training environment. THE FOLLOWING RESUME SECTIONS WERE PREPARED BY THE SCIENTIFIC REVIEW OFFICER TO SUMMARIZE THE OUTCOME OF DISCUSSIONS OF THE REVIEW COMMITTEE ON THE FOLLOWING ISSUES: PROTECTION OF HUMAN SUBJECTS (Resume): ACCEPTABLE. INCLUSION OF WOMEN PLAN (Resume): ACCEPTABLE. INCLUSION OF MINORITIES PLAN (Resume): ACCEPTABLE. INCLUSION OF CHILDREN PLAN (Resume): ACCEPTABLE.
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