Evaluation of the Roche COBAS® TaqMan® HIV-1 test for quantifying HIV-1 RNA in infected cells and lymphoid tissue

• Published in: Journal of virological methods Vol. 174; no. 1-2; pp. 69 - 76
• Main Authors: Gil, Cristina, Garcia, M. Teresa, Garcia, Felipe, Miró, José M., Agüero, Fernando, Alós, Lucía, Zamora, Laura, Capón, Alicia, Costa, Josep, Pumarola, Tomás, Gatell, José M.
• Format: Journal Article
• Language: English
• Published: Kidlington Elsevier B.V 01.06.2011; Elsevier
• Subjects: antiretroviral agents; Biological and medical sciences; Cells, Cultured; COBAS TaqMan HIV-1; detection limit; DNA; Fundamental and applied biological sciences. Psychology; HIV infections; HIV-1 - genetics; HIV-1 - isolation & purification; HIV-1 RNA quantitation; Human immunodeficiency virus 1; Humans; Lymphoid tissue; Microbiology; Nucleic Acid Amplification Techniques - methods; Palatine Tonsil - virology; patients; probit analysis; Reagent Kits, Diagnostic; Reproducibility of Results; RNA; RNA, Viral - genetics; RNA, Viral - isolation & purification; Sensitivity and Specificity; T-lymphocytes; T-Lymphocytes - virology; Techniques used in virology; tonsils; viral load; Viral Load - methods; Virology; HIV-1 RNA quantitation; COBAS TaqMan HIV-1; Lymphoid tissue; Immunopathology; Lymphoid cell; HIV-1 virus; Retroviridae; AIDS; Method; Immune deficiency; Lentivirus; Infection; Virus; Viral disease; Human immunodeficiency virus; Infected cell; Quantitative analysis
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2011.03.026

Lymphoid tissue is the main reservoir of HIV-1 in infected individuals. In this study the COBAS® TaqMan® HIV-1 test was evaluated for use with the High Pure System (HPS), for quantifying HIV-1 RNA in infected cells and lymphoid tissue specimens. Serial dilutions of 8E5–LAV1 infected T-cells into SUP-T1 cells and 44 tonsil specimens were examined. Some modifications of the test were required, such as the removal of residual DNA and the HIV-1 RNA output copies were adjusted to the sample input and expressed as HIV-1 RNA copies/μg of total RNA. The Roche COBAS® TaqMan HIV-1® (HPS) test proved to be a robust, sensitive, specific and reproducible method for quantifying HIV-1 RNA in infected cells and lymphoid tissue. Linearity and reproducibility were observed in serial dilutions of 8E5–LAV1 infected T-cells (R2>0.86). High reproducibility was found in clinical tonsil specimens (Wilcoxon test p>0.05). rDNase I treatment was essential to avoid false positives caused by residual HIV-1 DNA, mainly in tonsil specimens obtained from infected patients receiving effective antiretroviral treatment. Probit analysis determined the limit of detection as 22HIV-1 RNA copies/μg of total RNA. The Roche COBAS® TaqMan® HIV-1 (HPS) test thereby proved to be a helpful tool for measuring the HIV-1 viral load in infected cells and lymphoid tissue reservoirs.