title: Molecular typing for blood group antigens within 40 min by direct polymerase chain reaction from plasma or serum.
abstract: Determining blood group antigens by serological methods may be unreliable in certain situations, such as in patients after chronic or massive transfusion. Red cell genotyping offers a complementary approach, but current methods may take much longer than conventional serological typing, limiting their utility in urgent situations. To narrow this gap, we devised a rapid method using direct polymerase chain reaction (PCR) amplification while avoiding the DNA extraction step. DNA was amplified by PCR directly from plasma or serum of blood donors followed by a melting curve analysis in a capillary rapid-cycle PCR assay. We evaluated the single nucleotide polymorphisms underlying the clinically relevant Fya, Fyb, Jkaand Jkbantigens, with our analysis being completed within 40 min of receiving a plasma or serum sample. The positive predictive value was 100% and the negative predictive value at least 84%. Direct PCR with melting point analysis allowed faster red cell genotyping to predict blood group antigens than any previous molecular method. Our assay may be used as a screening tool with subsequent confirmatory testing, within the limitations of the false-negative rate. With fast turnaround times, the rapid-cycle PCR assay may eventually be developed and applied to red cell genotyping in the hospital setting.
authors.affiliation: DRK Blutspendedienst NSTOB, Institute Springe, Springe, Germany.
authors.forename: Franz F
authors.initials: FF
authors.lastname: Wagner
authors.affiliation: Department of Transfusion Medicine, NIH Clinical Center, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Willy A
authors.identifier: 0000-0002-1631-7198
authors.initials: WA
authors.lastname: Flegel
authors.affiliation: DRK Blutspendedienst NSTOB, Institute Springe, Springe, Germany.
authors.forename: Rita
authors.initials: R
authors.lastname: Bittner
authors.affiliation: DRK Blutspendedienst NSTOB, Institute Bremen-Oldenburg, Oldenburg, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Döscher
mesh_terms: D001782:Blood Donors; D001789:Blood Group Antigens / Q000097:blood*; D001788:Blood Grouping and Crossmatching; D005838:Genotype; D006801:Humans; D058889:Molecular Typing / Q000379:methods*; D010949:Plasma / Q000276:immunology; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D013997:Time Factors; D044366:Transition Temperature
chemical_list: D001789:Blood Group Antigens
keywords: blood groups; direct PCR; immunohematology; red cell genotyping; transfusion medicine