P045 Validation of BLDType for ABO/RHD/CCR5 typing of registry donors

Abstract

Aim To validate the BLDType Multiplex Typing Kit (ThermoFisher) for genotyping hematopoietic stem cell (HSC) registry donors for ABO, RHD and CCR5 using Fragment Analysis (FA). Methods Ninety-one archived genomic DNA samples (gDNAs) that had prior serology and/or ABO, RHD or CCR5 genotyping were typed using BLDType which employs fragment analysis using ABI 3130XL and Genemapper software 5 to distinguish A 1 , A 2 , B, O 1 , O 2 and O 3 alleles, detects deletion of RHD and CCR5 Δ 32. Fifty were typed for ABO and CCR5 alleles by restriction fragment length polymorphism (RFLP) and sequence-specific primer (SSP) PCR assays respectively. Twelve were genotyped for CCR5 by SSP and standard tube or automated red blood cell agglutination methods were used for ABO and RhD typing. Concordance rates were calculated for each sample subset as follows: 79 specimens with ABO and RhD phenotyping and 62 specimens with ABO by RFLP. Results and discussion Concordance between RhD serology and FA was 100% (79/79). Concordance between ABO serology and FA was 96% (76/79). The concordance between ABO RFLP and FA was 97% (60/62). The concordance between CCR5 genotyping by FA and SSP methods was 100% (91/91). The discrepancies encountered in this study were limited to ABO and were due to lack of serologic subtyping to distinguish A 1 from A 2 (N = 3) and a limitation of the RFLP which does not detect O 3 alleles (N = 2) or cannot resolve unusual banding patterns. The BLDType technique is user friendly and easy to implement. FA of 96 gDNAs can be completed in 8 h, and if automated and performed on a 3730XL, more than 1000 gDNAs can be tested per day. Conclusions The validation data presented here demonstrates that BLDType is a highly sensitive and specific typing tool. Incorporating BLDType along with HLA typing for registry donors would expedite the selection processes for both HLA-matched and ABO-compatible donors simultaneously.