Optimization of DNA collection card workflows on the RapidHIT™ ID instrument.

The RapidHIT™ ID System, enables CODIS eligible STR profile generation from reference samples in approximately 90 minutes. While initially optimized for buccal swabs, DNA collection cards, both treated (Nucleic-card™ and EasiCollect™) and untreated (Bode Buccal DNA Collector™), offer alternatives for collection and storage of reference samples. This study evaluated the performance of these cards using ACE GlobalFiler™ Express cartridges across a range of potentially impactful scenarios, including assessment of varying punch sizes, contamination control measures, treated versus untreated collection cards, sample placement within the cartridge, and impact of varying storage times on the collection cards. Results from cross-contamination testing confirmed no detectable contamination between samples, and a 6 mm punch size yielded the highest first-pass success rate and profile quality. Treated cards showed consistent performance during initial runs, regardless of whether they were processed wet or dry. For reruns, the EasiCollect™ cards on average produced higher overall signal when the wetting protocol was used. The untreated Bode card showed greater signal consistency when used dry, while wet treatment improved initial success rates. Proteinase K treatment enhanced DNA recovery from samples aged up to three years. Collectively, these findings provide practical, evidence-based guidelines for implementing DNA collection card-based workflows on the RapidHIT™ ID system, supporting its application across varied forensic and operational environments.