DETECTION AND DIRECT-SEQUENCE IDENTIFICATION OF BCR-ABL MESSENGER-RNAIN PH(-LEUKEMIA() CHRONIC MYELOID)

The reverse transcriptase-polymerase chain reaction (RT-PCR) for BCR-A BL mRNA is increasingly used to diagnose and monitor patients with Ph( +) chronic myeloid leukemia (CML). We investigated an alternative appr oach to detect BCR-ABL mRNA in CML in order to overcome some of the po tential drawbacks of RT-PCR. Nucleic acid sequence based amplification (NASBA) is a homogeneous, isothermal, in vitro process that provides the direct amplification of RNA, Peripheral blood from seven patients with Ph(+) CML and Ph(+) EM-2 cells were investigated by NASBA and RT- PCR. A nested set of four primers flanking the BCR-ABL junction was us ed in two serial NASBA reactions performed for 2 hours. The two method s were fully concordant for detection of transcripts with bcr3-ab12 an d bcr2-ab12 junctions. Ethidium bromide fluorescence with NASBA indica ted in repeated experiments that similar quantities of total RNA from patient material contained different amounts of BCR-ABL mRNA. The data suggest that direct amplification of RNA is suitable for identifying and monitoring patients with Ph(+) CML and may provide a means to quan tify BCR-ABL mRNA levels.