The frequency of serologically undetected human immunodeficiency virus type 1 (HIV-1) infection among a large cohort of seronegative intravenous drug users (IVDUs) was determined by polymerase chain reaction (PCR). In total, 2159 blood specimens were obtained from 945 seronegative IVDUs; of these, 182 had 1 specimen, 339 had 2, 397 had 3, and 27 had 4 semiannual specimens. No proviral DNA was detected in 2134 (98.8%) of the samples. Specimens from 7 persons (0.3%) were reactive by PCR. Within 6 months, all 5 of these 7 who returned for follow-up visits had seroconverted. Serum from 19 persons (0.9%) were equivocal by PCR analysis, that is, single primer pair amplification; 1 person seroconverted while others subsequently remained seronegative and nonreactive by PCR. The concordance between PCR and serology was 98.6%. It is concluded that immunosilent HIV-1 infection is uncommon and that serologic screening for HIV-1 antibodies is highly sensitive in this population.