High-throughput assessment of FMR1 and SNRPN methylation-based newborn screening using IsoPure and QIAcube HT systems

This study compared methylation-specific quantitative melt analysis of FMR1 and SNRPN methylation (mDNA) using automated bisulfite conversion by the magnetic-bead-based IsoPure and column-based QIAcube HT systems. Two bisulfite conversion methods were assessed on 3.2 mm punches from the same archival blood spots stored at room temperature for >10 years of individuals with FMR1 premutation (n = 20), fragile X syndrome (FXS, n = 20), or chromosome 15 imprinting disorders (n = 50) and freshly made blood spots from 184 newborns from the general population. Performance criteria were: (i) diagnostic sensitivity and specificity for the conditions screened; (ii) reaction failure rate; (iii) variability in mDNA between groups. Both methods showed 100% sensitivity and specificity for differentiating FXS and individual chromosome 15 imprinting disorders. IsoPure showed reaction failure rates of 0.365% for SNRPN and 0.74% for FMR1 compared to 19.34% and 2.56%, for QIAcube HT, respectively, with most failed reactions originating from archival blood spots. IsoPure showed lower variability in mDNA values in the neurotypical and condition-specific ranges. The IsoPure system showed superior performance especially on archival samples, with broader applications for screening and diagnostic testing requiring high-throughput mDNA analyses on materials of limited quantity and quality. Testing for a change to DNA known as methylation has been used by researchers and in medical practice to identify people affected with different diseases. We compared how well two procedures that employ robotics, work in a lab. We tested a small amount of blood soaked onto absorbent cards to detect this change. Blood on these cards was from people who either had fragile X syndrome or a chromosome 15 imprinting disorder or did not have these. Both procedures worked equally well on freshly made materials in identifying these conditions without making any mistakes. One of the procedures, however, worked better, on cards not recently made. These results provide new opportunities for automated testing of DNA to detect diseases.