In light of a recent report of 3 false-positive results of Epstein-Barr virus heterophile tests caused by HIV infection, we sought to assess the frequency of this occurrence. One hundred thirty-two positive heterophile antibody-tested serum samples were obtained from 2 tertiary care facilities in Boston to assess for HIV, and all tested negative for HIV plasma RNA. This study shows that false-positive results of heterophile tests are not frequently associated with primary HIV infection.