Abstract
Aim After transition to buccal swabs (SWs) for Be The Match Registry® recruitment activities in 2006, a study was initiated to evaluate the long-term stability of three sample types: frozen whole blood, whole blood spotted on filter paper (FP), and SW. Samples were analyzed for quality and quantity of DNA and accuracy of HLA typing. Data obtained from the first 5 years of the study are presented. Methods Fresh blood and SWs were collected from 30 volunteers with confirmed HLA typing. Fresh blood was stored as 1ml frozen aliquots and 75uL aliquots on FP. SWs and FP were stored at room temperature under controlled conditions. Samples were sent to 2 labs for testing at 6 time points (TP) immediately post collection and at one year storage intervals. Lab 1 performed high resolution HLA-A, B, C, DRB1 and DQB1 via SBT. Lab 2 performed intermediate resolution HLA-A, B, C and DRB1 via SSO and evaluated the quantity/quality of DNA within each sample. Results SSO and SBT HLA results were accurate at all loci through the 4 year TP. At 5 year TP, SSO results were 100% accurate, whereas SBT results were 99.4% accurate for FP and 97.3% for SWs. One FP and 4 SWs had amp failure, 5 SWs had allele drop-out and 1 SW had both. 19 FP and SW samples required repeat testing at multiple loci to obtain accurate HLA results. Sufficient DNA for HLA testing was extracted from all samples at all time points. DNA quality from frozen whole blood remained high thru year 5. DNA quality consistently decreased for the FP starting at the 3 year TP, 1 year TP for SWs. Conclusions Room temp storage of SWs and FP allows for DNA degradation, creating HLA testing inaccuracies and inefficiencies. Specifically, shorter amplicons were required to achieve accurate SBT results, though failures still persisted. DNA degradation did not affect SSO typing. Buccal swabs are a cost effective and efficient mechanism for registry DNA sample collection and storage, but shelf-life at room temperature is limited for Sanger SBT HLA typing method.
Published Version
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