Analytical validation of the amplification refractory mutation system polymerase chain reaction-capillary electrophoresis assay to diagnose spinal muscular atrophy.

Objectives: Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by homozygous deletion and compound heterozygous mutations in survival motor neuron 1 (), with severity tied to the copy number of survival motor neuron 2 (). This study aimed to develop a rapid and comprehensive method for the diagnosis of SMA.

Methods: A total of 292 children with clinically suspected SMA and 394 family members were detected by the amplification refractory mutation system polymerase chain reaction-capillary electrophoresis (ARMS-PCR-CE) method, which targeted 19 reported mutations, and the results were compared with those in multiplex ligation-dependent probe amplification (MLPA). Individuals with identified point mutations were further confirmed by long-range PCR and Sanger sequencing.

Results: A total of 202 children with SMA, 272 carriers, and 212 normal individuals were identified in this study. No difference was found in the R-value distribution of exons 7 and 8 in and among these cohorts, with coefficients of variation consistently below 0.08. To detect exon 7 and 8 copy numbers in and , the ARMS-PCR-CE results were concordant with those of MLPA. Approximately 4.95 % (10/202) of the study patients had compound heterozygous mutations.

Conclusions: The ARMS-PCR-CE assay is a comprehensive, rapid, and accurate diagnostic method for SMA that simultaneously detects copy numbers of exons 7 and 8 in /, as well as 19 point mutations in and 2 enhancers in . This approach can effectively reduce the time frame for diagnosis, facilitating early intervention and preventing birth defects.