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Updated: Oct 5, 2026
Multiplex PCR and Reverse Line Blot Hybridization Assay (mPCR/RLB)
Published on: August 6, 2011
Development of a Multiplex PCR to Detect the BCR-ABL1 Transcripts in Chronic Myeloid Leukaemia Patients in Indonesia
Delita Prihatni1,2, Nur Izzatun Nafsi1, Ernawati Ernawati1
1Department of Clinical Pathology, Faculty of Medicine/Dr. Hasan Sadikin General Hospital, Universitas Padjadjaran, Bandung, Indonesia.
Background:
There are limited commercial kits available for the identification of BCR-ABL1 gene fusion transcripts in chronic myeloid leukaemia (CML) patients. Most kits are based on RT-PCR to detect e14a2 and e13a2 transcripts as major breakpoint segments, but do not detect minor transcripts. Therefore, this study aimed to develop a multiplex PCR approach for BCR-ABL gene transcripts.
Methods:
Primers were designed to amplify four BCR-ABL1 transcripts from the cDNA reverse transcribed from 52 archived mRNAs from CML patients. The BCR-ABL1 transcripts were identified by multiplex PCR, and the assay performance was compared to Xpert® BCR-ABL Ultra (GeneXpert).
Results:
The multiplex TD-PCR successfully amplified four BCR-ABL1 transcripts, with detection limits of 10 fg/µL for e13a2 and e14a2 and 100 fg/µL for e1a2 and e19a2. Out of 52 archived RNA samples, 40 were confirmed positive. The most common transcript was e14a2, which was identified in 72.5% of the confirmed positive samples, followed by e13a2, identified in 25% of the positive samples. The e19a2 transcript was identified in 2.5% of the positive samples. No patients expressed the e1a2 transcript type or co-expressed both e14a2 and e13a2. The overall sensitivity and specificity of the multiplex PCR when compared to GeneXpert results were 97.56% (95% CI: 87.14-99.94%) and 100% (95% CI: 71.51-100%), respectively.
Conclusion:
The multiplex PCR assay developed to identify the four BCR-ABL1 transcripts can be implemented in CML diagnostic settings.
