NONRADIOACTIVE OLIGOTYPING FOR HLA-DR1-DRW10 USING POLYMERASE CHAIN-REACTION, DIGOXIGENIN-LABELED OLIGONUCLEOTIDES AND CHEMILUMINESCENCE DETECTION

We describe a rapid non-radioactive DNA typing of the serological types DR1-DRw10 using polymerase chain reaction (PCR)-amplified DNA and 15 sequence-specific oligonucleotides (SSO) which are labelled enzymatically at their 3' end with one digoxigenin (DIG). The hybridized SSOs were detected using anti-DIG alkaline phosphatase and Fab fragments and visualization was obtained with the chemiluminescent substate 3-(2'-spiroadamantan)-4-(3"-phosphoryloxy)-phenyl-1,2-dioxetan (AMPPD). The results were identical with those of the previously used 5-bromo-4-chloro-3-indolyl-phosphate (BCIP)/4-nitrobluetetrazolium chloride (NBT) system. The use of AMPPD is more rapid and allows the repeated rehybridization of the membrane-bound DNA.