The rapid and sensitive method for measuring reverse transcriptase (RT) activity developed in FY 89 was further improved in FY 90. The usefulness of detecting and monitoring HIV virus replication in susceptable T-cell (H9) and monocyte (U937) cell cultures was established for RT activity measured in cell lysates and culture medium. HIV p24 antigen expression is considered the most sensitive marker for HIV and is used to monitor cultures routinely. p24 antigen, however, is detectable in the absence of infectious virion. P24, therefore, is not a good candidate marker to access viral burden. RT activity, on the other hand, does accurately reflect the dynamic state of replication of infectious virion. The relative sensitivity of p24 antigen and RT activity measured by the rapid method for HIV were compared in freshly infected cells. Both markers were equally sensitive at detecting newly synthesized virus in cell lysates and culture supernatant. The quantitative relationship of RT activity and virus titer is being investigated. The quantitative, rapid, cell assay for infectious virion developed by Felber and Pavlakis was compared with RT and p24 antigen for early detection of virus replication. Samples of freshly infected cells and culture medium were tested for CAT activation with indicator cell lines containing stably Integrated copies of HIV-LTR-CAT. All three markers changed in parallel. CAT activation was greatly amplified, suggesting a basis for quantification. Results were presented at UCLA Conference on HIV and AIDS:Pathogenesis, Therapy and Vaccines (Abst.L 512). PCR technology represents a highly sensitive means of detecting virus-specific nucleic acids, although it is not yet available for routine monitoring of cell cultures. RT, p24, indicator CAT activity were found to be equally sensitive as p24 DNA determined by PCR analysis of HIV-infected cells. Results were presented at VI lnt. AIDS Mtg (Abst.FA 339).