Rapid Real-Time PCR Detection of HPdel Directly from Diluted Blood Samples

Abstract

Anhaptoglobinemic patients have been reported to experience severe anaphylactic reactions to transfusions due to the production of antihaptoglobin (anti-HP) antibodies(1)(2). Anhaptoglobinemia in patients homozygous for HP del , which is a deletion of an approximately 28-kb segment of chromosome 16 extending from the promoter region of the HP (haptoglobin) gene to exon 5 of HPR (haptoglobin-related protein), has been adequately characterized only recently(1). Use of a simple duplex PCR method has detected the HP del allele in East and Southeast Asian populations at frequencies of 1%–3% but this allele has not been detected in African, West and South Asian, and European populations(1)(3)(4). Thus, diagnosing HP del homozygosity prior to blood transfusion or the infusion of blood components into individuals from East and Southeast Asian populations is effective for preventing anaphylactoid shock due to anti-HP antibodies. We have developed a simple method that uses a 5′ nuclease real-time PCR assay (TaqMan; Applied Biosystems) to detect the HP del allele without having to isolate genomic DNA. The ethics committee of Kurume University School of Medicine approved this study protocol. To distinguish alleles, we performed real-time PCR assays that detect the 2 regions that encompass the HP del breakpoint and the 5′ …