Diagnosis of Toxoplasma gondii infection is difficult, especially in immunosuppressed people. The AIDS epidemic has increased the number of people at risk and increased the need for better diagnostic methods. We have compared three methods for detection of T. gondii parasitemia. Rabbits were infected subcutaneously with 104 T. gondii tachyzoites. Blood samples were obtained, and buffy coat or leukocyte fractions were prepared. We sought the T. gondii B1 gene by gene amplification by the polymerase chain reaction, and we sought viable T. gondii cells by inoculating fibroblast cell cultures and by mouse inoculation. Thirty-two blood samples were obtained from seven infected rabbits, and 18 were obtained from four control, uninfected rabbits. Parasitemia was detected in 20 of 32 samples (62%) from infected samples by mouse inoculation, 12 of 32 samples (37%) by gene amplification and detection, and 8 of 32 samples (25%) by cell culture. Mouse inoculation requires use of live animals and has a long turnaround time. Currently, cell culture is the least sensitive but most practical and widely available method for the detection of T. gondii parasitemia. Gene amplification and detection was more sensitive than cell culture and may become available in clinical laboratories as techniques are developed further and automated.