One-Pot Cascade Amplification Coupled with Lanthanide-Tagged ICP-MS for the Ultrasensitive Simultaneous Quantification of BCR-ABL Fusion Gene Subtypes

BCR-ABL fusion genes are gold-standard biomarkers for chronic myeloid leukemia (CML), but conventional detection methods suffer from cumbersome procedures, RNA preamplification needs, and an inability to quantify multiple subtypes simultaneously. Herein, we developed a one-pot enzyme-free cascade amplification system coupling toehold-mediated strand displacement reaction (TMSD) and hybridization chain reaction (HCR), combined with lanthanide-tagged inductively coupled plasma-mass spectrometry (ICP-MS), for the simultaneous quantification of four BCR-ABL subtypes (p190 e1a2, p210 e13a2, p210 e14a2, and p230 e19a2). The system uses tetrahedral DNAs (TDN1/TDN2) to optimize the probe orientation and amplification efficiency. Detection limits were 0.59, 0.82, 0.33, and 0.16 fM for the four subtypes, respectively. Validated in 70 clinical samples (40 CML patients, 30 healthy volunteers), it distinguished subtypes in bone marrow/peripheral blood, correlated well with qRT-PCR, and evaluated tyrosine kinase inhibitor (TKI) efficacy. This method avoids RNA reverse transcription and multitube reactions, providing a robust tool for CML diagnosis and targeted therapy monitoring.