HLA-DR,-DQ Typing By PCR Amplification With Sequence-specific Primers(PCR-SSP)

In most PCR-based tissue typing techniques the PCR amplification is followed by a post-amplification specificity step.ln typing by PCR amplification with sequence-specific primers(PCR-SSP),typing specificity is part of the amplification step,which make the technique almost as fast as serological tissue typing.In the study described here primers were designed for DR and DQ typing by PCR-SSP,i.e.identifying polymorphism corresponding to the serologically defined series DRI-DRI8,DQI-DQ9.This was achieved by performing 31 PCR reations per individual,20 for assigning DR1-DRI8,3 for the DR5I,DR52 and DR53 superspecificitys,and 8 for the DQ I -9.the reproducibility was 100% in 10 samples typed on two separated occasions.All samples could be typed successful ly,No false-positive or false-negative typing results were obtained.Amplification patterns segregated according to dominant Mendelian inheritance in four 2-generation families analysis .DNA preparation,PCR amplification and post-amplification pI~ocessing,including gel detection,documentation and interpretation,were performed in 4 hours .TD(touchdown)-PCR was designed for optimization of PCR amplification,the yield of amplified products were increased.In conclusion,PCR-SSP is an accurate typing technique with high sensitivitvspecificity and reproducibility.the method is rapid and inexpensive and is suited for routine clinical pratice.