Detection of Allergy-Related TPSAB1 Copy Number Variation by MALDI-TOF Nucleic Acid Mass Spectrometry.

BACKGROUND: Allergic diseases have been linked to genetic variations in TPSAB1 and TPSB2. This study aimed to develop a Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) method for simultaneous copy number detection of TPSAB1 and TPSB2. METHODS: We established a multiplex PCR-tandem MALDI-TOF MS assay for TPSAB1 and TPSB2 copy number analysis. To validate the method, 35 standard samples pre-validated by droplet digital PCR (ddPCR) were used to assess accuracy, sensitivity, repeatability, and stability. Additionally, we applied the method to 274 randomly selected population samples for clinical evaluation. RESULTS: The developed MALDI-TOF MS assay demonstrated high reliability, achieving 100% accuracy in standard samples. The detection limit for the standard template was 20 ng, with repeatability (CV: 8.41 - 13.60%) and stability (CV: 7.75 - 17.38%) within acceptable ranges. In the population cohort, the carrier rate for TPSAB1 copy number gain (> 2 copies) was 12.41%. CONCLUSIONS: The MALDI-TOF MS-based method is a robust, cost-effective, and high-throughput molecular diagnostic tool, particularly suited for differentiating highly homologous TPS genes. Its high accuracy and reliability make it a promising clinical assay for TPSAB1 and TPSB2 copy number analysis.