Olemukan R.E., Eller L.A., Ouma B.J., Etonu B., Erima S., Naluyima P., Kyabaggu D., Cox J.H., Sandberg J.K., Wabwire-Mangen F., Michael N.L., Robb M.L., De Souza M.S., Eller M.A.
Makerere University Walter Reed Project, Kampala, Uganda; U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States; International AIDS Vaccine Initiative, New York, NY, United States; Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 14186 Stockholm, Sweden; Armed Forces Research Institute of Medical Sciences, Bangkok, Thailand
Olemukan, R.E., Makerere University Walter Reed Project, Kampala, Uganda; Eller, L.A., Makerere University Walter Reed Project, Kampala, Uganda, U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States; Ouma, B.J., Makerere University Walter Reed Project, Kampala, Uganda; Etonu, B., Makerere University Walter Reed Project, Kampala, Uganda; Erima, S., Makerere University Walter Reed Project, Kampala, Uganda; Naluyima, P., Makerere University Walter Reed Project, Kampala, Uganda; Kyabaggu, D., Makerere University Walter Reed Project, Kampala, Uganda; Cox, J.H., International AIDS Vaccine Initiative, New York, NY, United States; Sandberg, J.K., Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 14186 Stockholm, Sweden; Wabwire-Mangen, F., Makerere University Walter Reed Project, Kampala, Uganda; Michael, N.L., U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States; Robb, M.L., U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States; De Souza, M.S., U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States, Armed Forces Research Institute of Medical Sciences, Bangkok, Thailand; Eller, M.A., Makerere University Walter Reed Project, Kampala, Uganda, U.S. Military HIV Research Program, 13 Taft Court, Rockville, MD 20850, United States, Department of Medicine, Karolinska Institutet, Karolinska University Hospital Huddinge, 14186 Stockholm, Sweden
Human immunodeficiency virus type 1 (HIV-1) vaccine and natural history studies are critically dependent on the ability to isolate, cryopreserve, and thaw peripheral blood mononuclear cell (PBMC) samples with a high level of quality and reproducibility. Here we characterize the yield, viability, phenotype, and function of PBMC from HIV-1-infected and uninfected Ugandans and describe measures to ascertain reproducibility and sample quality at the sites that perform cryopreservation. We have developed a comprehensive internal quality control program to monitor processing, including components of method validation. Quality indicators for real-time performance assessment included the time from venipuncture to cryopreservation, time for PBMC processing, yield of PBMC from whole blood, and viability of the PBMC before cryopreservation. Immune phenotype analysis indicated lowered B-cell frequencies following processing and cryopreservation for both HIV-1-infected and uninfected subjects (P < 0.007), but all other major lymphocyte subsets were unchanged. Long-term cryopreservation did not impact function, as unstimulated specimens exhibited low background and all specimens responded to staphylococcal enterotoxin B (SEB) by gamma interferon and interleukin-2 production, as measured by intracellular cytokine staining. Samples stored for more than 3 years did not decay with regard to yield or viability, regardless of HIV-1 infection status. These results demonstrate that it is possible to achieve the high level of quality necessary for vaccine trials and natural history studies in a resource-limited setting and provide strategies for laboratories to monitor PBMC processing performance. Copyright © 2010, American Society for Microbiology. All Rights Reserved.