RAPID CULTURE AND QUANTITATION OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1FROM PATIENT CELLS WITHOUT THE USE OF MITOGEN-STIMULATED DONOR CELLS

We report the development of a rapid, sensitive virus culture method f or direct quantitation of human immunodeficiency virus (HIV) in periph eral blood mononuclear cells (PBMCs). This new method involves culturi ng 10(7) PBMCs from HIV-seropositive persons in 10 ml of medium contai ning phorbol 12-myristate 13-acetate and interleukin-2. Both agents st imulate cell activation and hence viral replication. Cell-associated v irus and free virus are quantitated by a commercially available HIV p2 4 antigen capture enzyme immunoassay. Detection of cell-associated p24 antigen by flow cytometry was less sensitive than by the enzyme immun oassay. In this preliminary study, HIV was detected in 20 of 23 HIV-se ropositive patients and in none of the 11 HIV seronegative low-risk in dividuals. One HIV-seronegative person with Guillain-Barre syndrome fo llowing high-risk activity was found to be rapid-HIV-culture positive. The overall sensitivity and specificity were 87 and 100%, respectivel y. By comparing the quantity of virus produced in infected cells with the amount of virus produced in chronically infected U1 monocytes and ACH-2 lymphocytes stimulated with phorbol 12-myristate 13-acetate and interleukin-2, the approximate number of infected cells per sample is calculated. In the same patient specimens, quantitation of the number of HIV infected cells by the HIV rapid-culture method correlated with the results of the 21-day cell dilution coculture assay (correlation c oefficient r = 0.5; 95% confidence interval, 0.07 to 0.77). Advantages of the rapid HIV culture include no requirement for donor PBMCs or ch ange of media, shortened culture time, and the ability to detect p24 v iral antigen from cell-associated virus for quantitation of viral load .