Combined PCR-heteroduplex and PCR-SSCP analysis for matching of HLA-A, -B and -C allotypes in marrow transplantation.
We describe a method for rapid matching of HLA-A, -B and -C allotypes using simultaneous polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and heteroduplex analysis. Electrophoresis is performed at ambient temperature without requirements for buffer cooling. SSCP and heteroduplexes are revealed as discrete spatially separated band clusters. Using HLA-A, -B and -C locus-specific PCR primers, matching for alleles at these loci can be performed in 5 h. We tested 17 serologically matched patient-unrelated donor pairs and found considerable microheterogeneity at the DNA level. We propose that this technology has several advantages over conventional low-resolution typing methods and represents a potentially valuable screening method in unrelated donor selection.
Pursall
M C
MC
Department of Transplantation Sciences, University of Bristol, UK.
Clay
T M
TM
Bidwell
J L
JL
D001483:Base Sequence; D016026:Bone Marrow Transplantation; D017931:DNA Primers; D005091:Exons; D015234:HLA-A Antigens / Q000145:classification*; D015235:HLA-B Antigens / Q000145:classification*; D015236:HLA-C Antigens / Q000145:classification*; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D007126:Immunoglobulin Allotypes / Q000235:genetics*; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D018807:Polymorphism, Single-Stranded Conformational
D017931:DNA Primers; D015234:HLA-A Antigens; D015235:HLA-B Antigens; D015236:HLA-C Antigens; D007126:Immunoglobulin Allotypes; D009692:Nucleic Acid Heteroduplexes