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
| Autor: | Len Dally, Andrew Cooper, Simon Ogola, Pontiano Kaleebu, Ashwini Pujari, Peter Hayes, Mark Boaz, Patrick Mhlanga, Jill Gilmour, Jayashri Mahalingam, P. R. Narayanan, Josephine Birungi, Aloysius Semaganda, Madhuri Thakar, Abdul Mahmoud, Melanie Van Eeden, Tony Tarragona, Laura Seamons, Stephen Moore, Nilu Goonetilleke, Jackton Indangasi, Omu Anzala, Pattabiraman Sathyamoorthy, Bashir Farah, Gwynneth Stevens, Shadri Mishra, Paul Kato Kitandwe, V. D. Ramanathan, Josephine H. Cox, Dilbinder K. Gill |
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| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
Microbiology (medical)
Cell Survival Clinical Biochemistry Immunology Peripheral blood mononuclear cell Specimen Handling Interferon-gamma Immune system Immunity medicine Humans Immunology and Allergy Potency Interferon gamma Cells Cultured AIDS Vaccines Immunoassay Observer Variation medicine.diagnostic_test business.industry ELISPOT HIV Reproducibility of Results Vaccine Research Virology Leukocytes Mononuclear business medicine.drug |
| Popis: | The gamma interferon (IFN-γ) 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. PBMC recovery and viability after overnight resting and the IFN-γ 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-γ ELISPOT responses. These findings also illustrate the ability to standardize the IFN-γ 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. |
| Databáze: | OpenAIRE |
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