Virus load based on levels of functional reverse transcriptase (RT) was measured in plasma from 50 human immunodeficiency virus (HIV) type 1-infected persons, in 87 samples from 10 HIV-1 seroconversion panels, and in 100 uninfected persons by use of Amp-RT, an ultrasensitive RT assay. Of the 50 clinical samples, 38 (76%) were Amp-RT positive, while all uninfected controls were negative. Pearson's correlation coefficient of RNA and RT levels was .73 for all samples, .86 for seroconversion samples, and .49 for clinical samples. Calculated ratios of RT activity to virion RNA varied widely during both early and late stages of infection. Mean RT:RNA ratios in 8 seroconversion panels and in 12 (34.3%) of 35 individual clinical samples were significantly lower than the ratio for a reference virus. However, ratios were stable in individual seroconversions over time. These data demonstrate that RT activity can be used to quantitate plasma virus load and provide evidence of different levels of virion-associated RT among HIV-1-infected persons.