Rapid, direct, and qualitative method for the determination of the number of HIV-1-infected patient cells employing reactive oxygen intermediate generators

A method for the determination of the approximate number of HIV-1-infected patient cells in a patient sample, wherein said patient cells are subjected to direct stimulation in the absence of co-culture with donor cells, comprising the following steps:

• (a) obtaining peripheral blood mononuclear cells from a subject who is or may be infected with the HIV-1 virus and suspending said cells in culture media in a concentration of from about 0.5 to about 2.times.10.sup.6 /ml medium;
• (b) directly stimulating said peripheral blood mononuclear cells to activate expression of any HIV-1 virus present and corresponding p24 antigen production, in the absence of donor cell co-cultivation, by adding to said culture media phorbol-12-myristate-13-acetate in an amount of from 10 to 50 ng/ml media and interleukin-2 in an amount of from 6 to 20 units/ml media and incubating the resulting mixture for a period of from about 5 to about 7 days, at a temperature of from about 35.degree. to about 37.degree. C., a humidity of from about 90% to about 95%, and an atmospheric carbon dioxide concentration of from about 4% to about 5%;
• (c) separating the culture media following step (b) into cell and culture supernatant fractions and detecting any p24 antigen production in each of said cell and culture supernatant fractions;
• and
• (d) comparing the p24 antigen production of step (c) with standard values obtained from a culture comprising HIV-1 chronically infected cell lines of known proviral copy number directly stimulated as in step (b), wherein said standard values are obtained from a standard curve representing the amount of p24 antigen per number of cells from said HIV-1-infected cell lines;
• wherein said comparison results in the approximation of the number of HIV-1-infected patient cells in the sample.