Molecular methods for evaluation of virological status of nonhuman primates challenged with simian immunodeficiency or simian-human immunodeficiency viruses
| Autor: | Ranajit Pal, Lindsay Finke, Britany Bowen, Phillip D. Markham, Hye-Kyung Chung, Lauren Hudacik, Anthony D. Cristillo, Jessica Livesay, John J. Suschak, Eun Mi Lee, Lindsey Galmin |
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| Rok vydání: | 2010 |
| Předmět: |
Lymphoid Tissue
viruses Biology medicine.disease_cause Recombinant virus Sensitivity and Specificity Virus Plasma Proviruses Virology medicine Animals Self-Sustained Sequence Replication Immunodeficiency Temperature Reproducibility of Results RNA Viral Load Provirus Simian immunodeficiency virus medicine.disease biology.organism_classification Macaca mulatta NASBA DNA Viral Immunology Lentivirus HIV-1 Leukocytes Mononuclear RNA Viral Simian Immunodeficiency Virus |
| Zdroj: | Journal of Virological Methods. 163:287-294 |
| ISSN: | 0166-0934 |
| Popis: | Nonhuman primates represent a robust model to evaluate preclinical efficacy of HIV-1 vaccine and therapeutic strategies. Plasma and tissue viral RNA as well as tissue proviral DNA load are key parameters in assessing efficacy of vaccines and therapeutics against simian immunodeficiency virus (SIV) or simian-human immunodeficiency virus (SHIV) challenge. To quantitate SIV RNA in plasma and tissues, an isothermal nucleic acid sequence-based amplification (NASBA) method using real-time detection of amplified RNA with molecular beacons was developed. This assay has accuracy and reproducibility over seven orders of magnitude and has advantages over the electrochemiluminescence-based NASBA assay described previously, both in terms of higher throughput and sensitivity. Reproducibility and accuracy were also demonstrated for a TaqMan real-time PCR assay for quantitating proviral DNA load in PBMCs and lymphoid tissues. In infected macaques, the level of plasma viremia correlated with the tissue viral RNA but not always with proviral DNA loads. Further, animals with undetectable levels of viral RNA in plasma and proviral DNA in tissues, showed no sign of seroconversion and activation of Gag-specific CD8+ or CD4+ T cells in peripheral blood. These results suggest that simultaneous application of real-time NASBA and PCR assays provides quantitative evaluation of challenge outcome in macaques. |
| Databáze: | OpenAIRE |
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