Introduction of a real-time-based blood-group genotyping approach

Genotyping may be applied for rare blood group polymorphisms in a high-throughput mode as well for the molecular determination of blood groups due to unclear serological results.

We developed and validated a DNA typing method for the determination of KEL1/2, JK1/2, FY1/2, FY0, MNS1/2, MNS3/4, DO1/2, CO1/2 and LU1/2 alleles using a melting curve analysis downstream from a fully automated DNA extraction. All assays were validated in terms of specificity, sensitivity, assay variability and robustness. The usability was proven by a batch of 200 blood samples with partially known phenotype.

Assays for all blood groups were within the range of specificity (100%), assay variability and robustness (coefficient of variance < 3%). Genotypes of 200 random platelet donors were fully consistent with existing phenotype data. The obtained genotype distribution is in complete concordance with existing data for the European population underlined by a complete absence of CO2 homozygous donors and the FY0 allele among the cohort.

We introduce an approach for blood group genotyping of particular samples or gene loci in glass capillary format and for medium-throughput analysis in 96/384-well format. The advantages of this real-time polymerase chain reaction method are its automation potential, the flexibility regarding hardware and the rapid cycling time.