A tetrazolium-based colorimetric assay for quantification of HIV-1-induced cytopathogenicity in monocyte-macrophages exposed to macrophage-colony-stimulating factor.

A sensitive assay was developed for in vitro evaluation of anti-HIV agents in monocyte-macrophage cells (M/M) (a crucial target of HIV in the body). Monocyte-macrophage cells are usually poorly sensitive to the cytopathic effect induced by HIV. However, when fresh adherent monocyte-macrophage cells are cultured at relatively high density in the presence of macrophage-colony stimulating factor (M-CSF), they undergo cytolysis and die in 2-3 weeks. HIV-mediated cell-killing can thus be assessed with a method based on the reduction of the yellow colored 3-(4-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) by metabolically active cells to a blue formazan, which can be measured spectrophotometrically. HIV-mediated cytopathic effect of M-CSF-exposed monocyte-macrophage cells was consistently achieved in all experiments performed under the conditions described herein. Anti-HIV activity of zidovudine (AZT) was also comparatively evaluated in M-CSF- and normal monocyte-macrophage cells both using the MTT assay and by measuring HIV-p24 antigen production in supernatants of monocyte-macrophage cells cultures, and similar results obtained with both methods. These results support the use of this colorimetric assay for broad screening of anti-HIV agents in monocyte-macrophage cells.