Development and implementation of an international proficiency testing program for a neutralizing antibody assay for HIV-1 in TZM-bl cells

• Published in: Journal of immunological methods Vol. 375; no. 1-2; pp. 57 - 67
• Main Authors: Todd, Christopher A., Greene, Kelli M., Yu, Xuesong, Ozaki, Daniel A., Gao, Hongmei, Huang, Yunda, Wang, Maggie, Li, Gary, Brown, Ronald, Wood, Blake, D'Souza, M. Patricia, Gilbert, Peter, Montefiori, David C., Sarzotti-Kelsoe, Marcella
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 31-01-2012; Elsevier
• Subjects: AIDS Vaccines - immunology; Antibodies, Neutralizing - analysis; Antibodies, Neutralizing - immunology; Antibody; Assay; Biological and medical sciences; Cell Line, Tumor; clinical trials; Fundamental and applied biological sciences. Psychology; Fundamental immunology; HIV; HIV Antibodies - analysis; HIV Antibodies - immunology; HIV Infections - immunology; HIV Infections - virology; HIV-1 - immunology; Human immunodeficiency virus 1; Humans; Laboratory Proficiency Testing - standards; luciferase; Microbiology; Molecular immunology; neutralization; Neutralization Tests - standards; Neutralizing; neutralizing antibodies; Proficiency; Quality Control; Reference Standards; reporter genes; risk; Techniques; Techniques used in virology; TZM-bl; vaccines; Virology; Assay; Proficiency; HIV; Antibody; Neutralizing; TZM-bl; HIV-1 virus; Retroviridae; Lentivirus; Neutralizing antibody; Immunological method; Virus; Human immunodeficiency virus
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2011.09.007

Recent advances in assay technology have led to major improvements in how HIV-1 neutralizing antibodies are measured. A luciferase reporter gene assay performed in TZM-bl (JC53bl-13) cells has been optimized and validated. Because this assay has been adopted by multiple laboratories worldwide, an external proficiency testing program was developed to ensure data equivalency across laboratories performing this neutralizing antibody assay for HIV/AIDS vaccine clinical trials.
The program was optimized by conducting three independent rounds of testing, with an increased level of stringency from the first to third round. Results from the participating domestic and international laboratories improved each round as factors that contributed to inter-assay variability were identified and minimized. Key contributors to increased agreement were experience among laboratories and standardization of reagents.
A statistical qualification rule was developed using a simulation procedure based on the three optimization rounds of testing, where a laboratory qualifies if at least 25 of the 30 ID50 values lie within the acceptance ranges. This ensures no more than a 20% risk that a participating laboratory fails to qualify when it should, as defined by the simulation procedure. Five experienced reference laboratories were identified and tested a series of standardized reagents to derive the acceptance ranges for pass–fail criteria. This Standardized Proficiency Testing Program is the first available for the evaluation and documentation of assay equivalency for laboratories performing HIV-1 neutralizing antibody assays and may provide guidance for the development of future proficiency testing programs for other assay platforms.