Stocks of HIV-1 strains JR-CSF and NL4-3 were produced by plasmid transfection of 293T cells, as previously described.60 HIV neutralization activity was measured for serial dilutions of the supernatants from J3 or A6-edited B cell lines using the TZM-bl assay, as previously described.61 (link) Luminescence was detected with the britelite plus kit (Perkin-Elmer) and measured with a Mithras LB 940 plate reader (Berthold Technologies). Control J3 and A6 HCAbs were generated from transfected 293T cells. Input amounts of each virus were titrated to give roughly 10-20 times higher luminescence than background in control wells receiving no virus or antibodies.
Protocol detail
Quantification of HIV-1 Neutralization
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293t cells
Antibodies
Assay
Cell lines
Dilutions
Hiv 1
Luminescence
Plasmid
Strains
Transfection
Virus
Corresponding Organization : University of Southern California
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Variable analysis
independent variables
- Stocks of HIV-1 strains JR-CSF and NL4-3
- Serial dilutions of the supernatants from J3 or A6-edited B cell lines
- HIV neutralization activity measured using the TZM-bl assay
- Input amounts of each virus were titrated to give roughly 10-20 times higher luminescence than background in control wells receiving no virus or antibodies
- Positive control: Control J3 and A6 HCAbs generated from transfected 293T cells
- Negative control: Control wells receiving no virus or antibodies
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