The detection of residual acute lymphoblastic leukemia cells with immunologic methods and polymerase chain reaction:...

In this study we applied double color immunofluorescence analysis and polymerase chain reaction (PCR) amplification of rearranged TCR delta genes for detecting residual leukemia in the posttreatment bone marrow (BM) samples taken from four patients in morphological remission. In three of these patients (nos. 1-3; T-ALL) a combination of CD3 and anti-TdT antibodies (Abs) was used to identify residual blasts while in patient 4 (B lineage ALL) the combination CD13/TdT served to detect residual disease. Two rounds of PCR primed by nested amplimers were carried out to prepare clonospecific probes from presentation DNA and to investigate the follow-up samples. In patients 1 and 2 no cCD3+/TdT+ cells were seen posttreatment, but PCR amplification of the TCR V delta 1-D-J delta 1 region revealed residual disease in both patients. Patient 1 underwent allogeneic BM transplant (BMT) 8 months after diagnosis and is well 3 months post-BMT while patient 2 relapsed 12 months after presentation. In patient 3 the remission samples investigated 2 and 3 months after diagnosis did not contain cCD3+/TdT+ cells, but in the sample collected at 4 months a few such cells (0.0001-0.001%) could be detected. In the same sample, PCR amplification of the TCR V delta 2-D-J delta 1 region indicated the presence of 10(-4)-10(-3) residual leukemic cells. These findings predicted full morphological relapse which occurred 2 months later. In patient 4 CD13/TdT double positive cells were clearly seen 2 and 3 months after presentation. PCR amplification of the V delta 2-D delta 3 recombination also revealed residual blasts when applied to one of such "remission" samples. After further remission induction treatment, no immunologic evidence of residual disease was detected. This patient received an allogeneic BMT 8 months after diagnosis and is disease free 4 months after BMT. Our data indicate that both double color immunofluorescence and PCR analysis offer powerful tools to study residual leukemia and highlight the advantages as well as the potential limitations of each technique.