Viral Neutralization Assay for HIV-1 Env

Viral neutralization by trial participant serum samples were measured using single-round HIV-1 Env pseudovirus infection of TZM-bl target cells. Env-pseudotyped viruses were generated by transfection of 293T/17 cells with optimized ratios of envelope-expressing plasmid and backbone vector (pSG3DEnv). A panel of three viruses was tested for neutralizing activity spanning a range of 80% inhibitory concentrations (IC₈₀) for bnMAb VRC01. Negative and positive controls were included in all assays. The tested viruses were Q23.17 (subtype A), PVO.04 (subtype B), and MW965.26 (subtype C). Serum neutralization assays were performed in 384-well plates using a Beckman Biomek liquid handling system, as previously described [33 (link)]. Experimental results were displayed as the serum dilution that produced 80% neutralization (ID₈₀) against the viruses tested. Predicted ID₈₀ values were calculated based on the concentration of VRC01LS present in the sera and the established inhibitory concentration (IC₈₀) of the antibody against each virus.

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Gaudinski M.R., Coates E.E., Houser K.V., Chen G.L., Yamshchikov G., Saunders J.G., Holman L.A., Gordon I., Plummer S., Hendel C.S., Conan-Cibotti M., Lorenzo M.G., Sitar S., Carlton K., Laurencot C., Bailer R.T., Narpala S., McDermott A.B., Namboodiri A.M., Pandey J.P., Schwartz R.M., Hu Z., Koup R.A., Capparelli E., Graham B.S., Mascola J.R, & Ledgerwood J.E. (2018). Safety and pharmacokinetics of the Fc-modified HIV-1 human monoclonal antibody VRC01LS: A Phase 1 open-label clinical trial in healthy adults. PLoS Medicine, 15(1), e1002493.

Publication 2018

liquid handling system

293t cells Assays Backbone Cells Dilution Hiv 1 Infection Inhibitory Plasmid Pseudotyped viruses Sera Transfection Vector Virus antibody Viruses

Corresponding Organization : National Institutes of Health

Other organizations : Medical University of South Carolina, University of California, San Diego

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Variable analysis

independent variables

Serum samples from trial participants

dependent variables

Viral neutralization by trial participant serum samples
Serum dilution that produced 80% neutralization (ID80) against the viruses tested

control variables

Env-pseudotyped viruses generated by transfection of 293T/17 cells with optimized ratios of envelope-expressing plasmid and backbone vector (pSG3DEnv)
Panel of three viruses tested for neutralizing activity: Q23.17 (subtype A), PVO.04 (subtype B), and MW965.26 (subtype C)
Negative and positive controls included in all assays

controls

Negative controls
Positive controls

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