Detection and direct sequence identification of BCR-ABL mRNA in Ph+ chronic myeloid leukemia.

The reverse transcriptase-polymerase chain reaction (RT-PCR) for BCR-ABL mRNA is increasingly used to diagnose and monitor patients with Ph+ chronic myeloid leukemia (CML). We investigated an alternative approach to detect BCR-ABL mRNA in CML in order to overcome some of the potential 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 used in two serial NASBA reactions performed for 2 hours. The two methods were fully concordant for detection of transcripts with bcr3-abl2 and bcr2-abl2 junctions. Ethidium bromide fluorescence with NASBA indicated 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 quantify BCR-ABL mRNA levels.