Variability in the levels of PML-RAR alpha fusion transcripts detected by laboratories participating in an external quality control program using several reverse transcription polymerase chain reaction protocols

Background and Objectives. The detection of PML-RAR by reverse transcriptio n (RT) polymerase chain reaction (PCR) in acute promyelocytic leukemia (APL ) patients who are in hematologic remission influences therapeutic decision making in several trials. In the light of this, the Spanish group has rece ntly designed an external quality assessment program (EQAP) of RT-PCR detec tion of PML-RAR, which includes a study of sensitivity of the participating laboratories. Design and Methods. Eighteen laboratories were Involved in the program. Ten laboratories followed the method of Biondi et al.,(4) 5 employed that of B orrow et al.(10) and the 3 remaining used other protocols. The sensitivity was studied in five rounds of quality control. The first two shipments cons isted of dilutions of NB4 RNA into non-Apt. RNA. The third round consisted of serial dilutions of the NB4 cell line into HL60 cells. The fourth and fi ve rounds consisted of plasmid dilutions containing the bcr1 and bcr3 PML-R AR isoforms. Results. The results showed that the distinct methods allow detection of th e PML-RAR hybrid up to a dilution of 10(-4), and exceptionally, up to 10(-5 ). The laboratories following the method of Biondi et al. usually detected the 10(-3) dilution and less frequently the 10-4 one, whereas those using o ther methods usually detected PML-RAR transcript in the 10(-4) dilution, an d less commonly in the 10-5 dilution. However, each of the PCR methods used by EQAP participating laboratories successfully detected at least 50 copie s of PML-RAR alpha fusion transcript in plasmid dilution controls. Interpretation and Conclusions. The results point to heterogeneous sensitiv ity amongst participating laboratories. This may reflect differences in met hodology, although variations in sample quality may also account for discre pant findings. (C) 2001, Ferrata Storti Foundation.