Chimerism monitoring using biallelic single nucleotide or insertion/deletion polymorphisms: How many markers to screen?

ABSTRACT

BACKGROUND/AIMS: Chimerism monitoring by means of high-throughput sequencing or quantitative PCR of biallelic single nucleotide and insertion/deletion polymorphisms has shown potential for improved patient care when compared to the gold standard capillary electrophoresis assays. When designing chimerism assays the number of markers to screen needs consideration it determines the informativity rate and accuracy of the assay, but screening too many markers increases the assay's cost and complexity. The minimal number of biallelic markers to screen is currently unstudied. MATERIALS/METHODS: A simulation framework accounting for marker minor allele frequencies, the number of markers screened, marker allelic constellations and donor-recipient relatedness was constructed. The framework was validated through analysis of 324 clinical samples. RESULTS: Empirical clinical data confirm the validity of the simulation framework. With guidelines suggesting to monitor at least three informative markers, we demonstrate that, for optimized assays, at least 40 biallelic markers need to be screened to achieve enough informative markers in over 99% of cases. We propose and discuss several assay optimization strategies. CONCLUSION: Currently used chimerism assays often screen too little or too many markers, leaving room for optimization. Through support of the simulation framework here introduced and validated, more informative, cost-effective chimerism assays can be designed.