Flow cytometric assay for HIV-1 whole virion binding: virus inactivated with aldrithiol-2 has binding properties comparable to native virions

HIV-1 virions selectively incorporate host cell derived proteins in the process of budding. In particular, MHC Class II molecules (HLA-DR) are readily incorporated into virions produced from HLA-DR+ cells, a fact that can be exploited to detect binding to target cells. Under suitable conditions, if HLA-DR+ virions produced from HLA-DR+ cell lines are incubated with CD4+, HLA-DR- target cells, acquisition of HLA-DR+ reactivity by the target cells can be used as a criterion of viron binding. We screened numerous cell lines to identify candidate CD4+, HLA-DR- target cells for such an assay, using both crude (neat infected cell supernatant) and purified (concentrated, sucrose banded) HLA-DR+ virions, controlling for the presence of non-viral microvesicles in the virus preparations. Among the CD4+, HLA-DR- cell lines identified, we observed varying degrees of both specific (blockable by target cell pre-incubation with anti-Leu 3a) and non-specific (not anti-Leu 3a mAb blockable) binding, that did not correlate with CXCR4 co-receptor expression. The A3.01 cell line was selected as the line showing the greatest degree of specific binding. Virus binding was concentration dependent, blockable with anti-Leu 3a and associated with loss anti-Leu 3a, but not OKT4 mAb reactivity. In contrast to virions inactivated with heat or formalin, virions inactivated with aldrithiol-2 showed binding properties comparable to native virus, indicating that this mode of inactivation, which eliminates infectivity, preserves the conformational and functional integrity of virion surface proteins.

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