Aim: To develop a fully automated hybrid capture-based target enrichment script for the One Lambda HybriType HLA Plus assay on a bench-top digital microfluidics device.
Methods: Human leukocyte antigen (HLA) typing can be challenging due to the high level of genetic diversity within the HLA genes. Hybrid capture-based target enrichment paired with next-generation sequencing (NGS) provides a robust approach by enabling selective isolation and sequencing of HLA regions. The One Lambda HybriType assay is specifically designed for comprehensive genotyping, covering 19 loci—including 11 HLA genes and 7 extended MHC loci—as well as ABO gene. Automation was implemented using the MIRO CANVAS system, which utilizes digital microfluidics within single-use cartridges. With integrated thermal cycling and magnetic bead handling, the system enables fully walkaway automation, completing the 3-hour protocol with less than 15 minutes of hands-on time. Automated results were compared to the manual workflow to confirm high-resolution HLA typing with consistency, precision, and efficiency.
Results: Using the same set of 24 DNA samples, we performed three automated and three manual hybrid capture replicates (six runs total). All runs performed on the MIRO CANVAS platform met the predefined quality criteria for read depth, allele balance, and minimum coverage. In the automated runs, the on-target rate was 56%, duplication rate was 4.4%, and the average insert size was 512 bp. In the manual runs, the on-target rate was 75%, duplication rate was 8.8%, and the average insert size was 527 bp. Allele concordance was 100% for both the automated and manual workflows.
Conclusion: Automation of the HybriType assay using the MIRO CANVAS system produced high-quality results with minimal hands-on time. The system’s microfluidic channels and electrowetting technology enabled precise droplet control, reducing user error and improving consistency. Additionally, an onboard touchscreen guided users through setup, ensuring reliable protocol adherence. Overall, automation significantly reduces hands-on time while ensuring high-quality, consistent results, making the automated HybriType assay an invaluable tool for laboratories requiring detailed, reliable genomic insights.
