Quantifying HIV-1 RNA Induction

Total CD4⁺ T cells were isolated from PBMCs (Stem Cell) and cultured (3–6 replicates of 1 million cells/well) in phenol red–free R10 media with or without phorbol 12-myristate 13-acetate (PMA)/ionomycin (Iono) for 7 days in the presence of 300 mM of efavirenz and raltegravir as previously described [21 (link)]. Total induced HIV-1 RNA was quantified in culture supernatants (pooled from replicate wells and tested in duplicate) by qPCR at pre-entry, week 6, and week 12.

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Riddler S.A., Zheng L., Durand C.M., Ritz J., Koup R.A., Ledgerwood J., Bailer R.T., Koletar S.L., Eron J.J., Keefer M.C., Macatangay B.J., Cyktor J.C, & Mellors J.W. (2018). Randomized Clinical Trial to Assess the Impact of the Broadly Neutralizing HIV-1 Monoclonal Antibody VRC01 on HIV-1 Persistence in Individuals on Effective ART. Open Forum Infectious Diseases, 5(10), ofy242.

Publication 2018

PBMCs

Cd4 t cells Cells Efavirenz Hiv 1 Ionomycin Phorbol myristate acetate Raltegravir Replicate Stem cell

Corresponding Organization : University of Pittsburgh

Other organizations : Cancer Research And Biostatistics, Johns Hopkins University, National Institute of Allergy and Infectious Diseases, National Institutes of Health, The Ohio State University, University of North Carolina at Chapel Hill, University of Rochester

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Variable analysis

independent variables

Presence or absence of phorbol 12-myristate 13-acetate (PMA)/ionomycin (Iono)

dependent variables

Total induced HIV-1 RNA quantified in culture supernatants

control variables

Total CD4+ T cells isolated from PBMCs (Stem Cell)
Cells cultured in phenol red–free R10 media
Presence of 300 mM of efavirenz and raltegravir
Cells cultured for 7 days

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