Rapid, single-subject genotyping to predict red blood cell antigen expression

Area of Science:

• Transfusion Medicine
• Genetics
• Molecular Biology

Background:

• Genotyping predicts red blood cell (RBC) antigen expression when antisera are scarce.
• It aids in determining phenotypes for highly transfused patients.
• Genotyping stored DNA is an option when fresh blood samples are unavailable.

Purpose of the Study:

• To evaluate a sequence-specific primer (SSP) genotyping system for blood group alleles.
• To assess the suitability of SSP for rapid testing of small sample batches.
• To determine the efficacy of using stored whole blood for genotyping.

Main Methods:

• Genomic DNA was extracted from fresh and 1- to 2-week-old stored blood (n=20).
• ABO, RHD, and RHCE genotyping was performed using SSPs.
• Amplicons were analyzed via gel electrophoresis and a microfluidic on-chip electrophoresis system.

Main Results:

• SSP and gel electrophoresis correctly typed all fresh and stored blood samples for ABO, RHD, and RHCE.
• On-chip electrophoresis provided accurate typing for most samples, with minor discrepancies in RHCE.
• On-chip electrophoresis used 10% of the DNA and was 4x faster than gel electrophoresis, with improved amplicon visualization.

Conclusions:

• SSP genotyping is feasible for ABO and RH typing using fresh and stored blood samples.
• On-chip electrophoresis is a rapid, DNA-efficient, and user-friendly method for blood group genotyping.
• While faster, on-chip electrophoresis showed slightly increased variability in test reliability compared to gel electrophoresis.