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Development of Real-time PCR and Melting Curve Analysis for the Rapid Detection of DEL with RHD (c.1222T>C) or RHD (c.1227G>A) by Yun Ji Hong, Hyungsuk Kim, Eun Young Song, Duck Cho, Kyoung Un Park, Kyou-Sup Han

Background: DEL, a variant of RhD, is difficult to detect in routine blood bank testing owing to its extremely low levels of D antigen expression. However, DEL is capable of alloimmunizing a patient when transfused into RhDnegative individuals.
Methods: In this study, we developed real-time PCR and melting curve analysis for the rapid detection of the DEL phenotype.
Results: Of the 325 serologically RhD-negative individuals involved in the study, 56 (17.2%) had melting temperatures distinguishable from complete RHD absence as follows: 53 RHD (c.1227G>A) DEL specimens had a plateau at 54 - 56°C and a peak at 61.95°C, while 3 RHD (c.1222T>C) DEL had a melting temperature of 62.62°C. All DEL results were identical to those obtained by multiplex single-base primer extension reactions.
Conclusions: The rapid DEL genotyping method developed in this study will be useful for screening DEL in serologically RhD-negative donors and preventing the alloimmunization of RhD-negative individuals by DEL.

DOI: 10.7754/Clin.Lab.2016.160320