NEUTRALIZING ANTIBODY EVALUATION INFECTIVITY REDUCTION ASSAY

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Tony Rook
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NEUTRALIZING ANTIBODY EVALUATION INFECTIVITY REDUCTION ASSAY

Please find the link for the following protocol:

NEUTRALIZING ANTIBODY EVALUATION INFECTIVITY REDUCTION ASSAY

http://aactg.s-3.com/pub/download/imm/imhivneu.pdf

Brief Description:

The HIV-1 neutralizing antibody assessment by the infectivity reduction assay estimates the reduction of the number of infectious units per milliliter (IUPML) in 1 mL of viral stock dilutions when treated with HIVIG or patient serum. This assay is performed in a 24-well plate and is based on the ACTG consensus plasma quantitative culture assay, using six 5-fold dilutions of viral isolate neutralized by HIVIG, serum or plasma. A 1:125 final concentration of HIVIG is added to each of the
dilutions. When autologous serum is used, 1:20 or 1:40 dilution of serum is preferred. Each sample of viral dilution is cocultured with PHA-stimulated normal donor PBMC for 14 days. The supernatant for each individual well is assayed for HIV-1 p24 antigen as a determination of viral growth. The antigen results from each well can be evaluated by a software program to determine the IUPML in the presence of HIVIG, patient serum or normal human serum (control) according to the ACTG Virology Manual. The difference between the IUPML would be used to assess neutralizing capabilities of the test specimen. Based on preliminary studies, assay variation is expected to be 0.7 log.