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    Concordant Proficiency in Measurement of T-Cell Immunity in Human Immunodeficiency Virus Vaccine Clinical Trials by Peripheral Blood Mononuclear Cell and Enzyme-Linked Immunospot Assays in Laboratories from Three Continents

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    The final, definitive version of this paper has been published in the Clinical and vaccine immunology, Feb. 2009, Vol. 16 No. 2 p. 147–155. 1556. DOI:10.1128/CVI.00326-08, published by the American Society for Microbiology. All rights reserved. (954.9Kb)
    Date
    2009-02
    Author
    Boaz, Mark J.
    Hayes, Peter
    Tarragona, Tony
    Seamons, Laura
    Cooper, Andrew
    Birungi, Josephine
    Kitandwe, Paul
    Semaganda, Aloysius
    Kaleebu, Pontiano
    Stevens, Gwynneth
    Anzala, Omu
    Farah, Bashir
    Ogola, Simon
    Indangasi, Jackton
    Mhlanga, Patrick
    Eeden, Melanie Van
    Thakar, Madhuri
    Pujari, Ashwini
    Mishra, Shadri
    Goonetilleke, Nilu
    Moore, Stephen
    Mahmoud, Abdul
    Sathyamoorthy, Pattabiraman
    Mahalingam, Jayashri
    Narayanan, Paranji R.
    Ramanathan, Vadakkuppattu D.
    Cox, Josephine H.
    Dally, Len
    Gill, Dilbinder K.
    Gilmour, Jill
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    Abstract
    The gamma interferon (IFN-_) enzyme-linked immunospot (ELISPOT) assay is used routinely to evaluate the potency of human immunodeficiency virus (HIV) vaccine candidates and other vaccine candidates. In order to compare candidates and pool data from multiple trial laboratories, validated standardized methods must be applied across laboratories. Proficiency panels are a key part of a comprehensive quality assurance program to monitor inter- and intralaboratory performance, as well as assay performance, over time. Seven International AIDS Vaccine Initiative-sponsored trial sites participated in the proficiency panels described in this study. At each laboratory, two operators independently processed identical sample sets consisting of frozen peripheral blood mononuclear cell (PBMC) samples from different donors by using four blind stimuli. PBMCM recovery and viability after overnight resting and the IFN-_ ELISPOT assay performance were assessed. All sites demonstrated good performance in PBMC thawing and resting, with a median recovery of 78% and median viability of 95%. The laboratories were able to detect similar antigen-specific T-cell responses, ranging from 50 to >3,000 spot-forming cells per million PBMC. An approximate range of a half log in results from operators within or across sites was seen in comparisons of antigen-specific responses. Consistently low background responses were seen in all laboratories. The results of these proficiency panels demonstrate the ability of seven laboratories, located across three continents, to process PBMC samples and to rank volunteers with differential magnitudes of IFN-_ ELISPOT responses. These findings also illustrate the ability to standardize the IFN-_ ELISPOT assay across multiple laboratories when common training methods, reagents such as fetal calf serum, and standard operating procedures are adopted. These results are encouraging for laboratories that are using cell-based immunology assays to test HIV vaccines and other vaccines.
    Use this URI to cite this item:
    https://hdl.handle.net/20.500.11951/326
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