J Yankowitz1, S Li, J C Murray. 1. Department of Obstetrics and Gynecology, University of Iowa School of Medicine, Iowa City, USA.
Abstract
OBJECTIVE: To evaluate the accuracy of polymerase chain reaction (PCR) amplification of a portion of the RhD gene by testing a large number of DNA samples derived from individuals whose RhD status was established by the standard serologic method. METHODS: Seven hundred sixty-five samples were obtained from two sources: subjects taking part in studies at the University of Iowa Hospitals and Clinics (n = 107), and Centre d'Etude du Polymorphisme Humain (CEPH) families used for studies of genetic variation (n = 658). Deoxyribonucleic acid was extracted from blood samples of University of Iowa volunteers and from CEPH families by standard techniques. With few modifications, published primers, reaction and electrophoresis conditions, which yield a 1200-bp fragment in all individuals and a 600-bp fragment in RhD-positive individuals, were used. RESULTS: By standard serologic techniques, we identified samples from 632 RhD-positive and 133 RhD-negative individuals. Two (both from CEPH) of the 632 RhD-positive individuals were characterized as RhD-negative by PCR. Seven of the 133 RhD-negative samples were judged to be RhD-positive by PCR because of the presence of a light 600-bp band. Despite repeated attempts, no bands or DNA were identified in one RhD-negative sample. Thus, the sensitivity of RhD typing by PCR was 99.7%, the specificity 94.0%. CONCLUSION: Based on our data, it would appear that the use of PCR to establish RhD type can be introduced cautiously into current management schemes in the evaluation of RhD sensitization.
OBJECTIVE: To evaluate the accuracy of polymerase chain reaction (PCR) amplification of a portion of the RhD gene by testing a large number of DNA samples derived from individuals whose RhD status was established by the standard serologic method. METHODS: Seven hundred sixty-five samples were obtained from two sources: subjects taking part in studies at the University of Iowa Hospitals and Clinics (n = 107), and Centre d'Etude du Polymorphisme Humain (CEPH) families used for studies of genetic variation (n = 658). Deoxyribonucleic acid was extracted from blood samples of University of Iowa volunteers and from CEPH families by standard techniques. With few modifications, published primers, reaction and electrophoresis conditions, which yield a 1200-bp fragment in all individuals and a 600-bp fragment in RhD-positive individuals, were used. RESULTS: By standard serologic techniques, we identified samples from 632 RhD-positive and 133 RhD-negative individuals. Two (both from CEPH) of the 632 RhD-positive individuals were characterized as RhD-negative by PCR. Seven of the 133 RhD-negative samples were judged to be RhD-positive by PCR because of the presence of a light 600-bp band. Despite repeated attempts, no bands or DNA were identified in one RhD-negative sample. Thus, the sensitivity of RhD typing by PCR was 99.7%, the specificity 94.0%. CONCLUSION: Based on our data, it would appear that the use of PCR to establish RhD type can be introduced cautiously into current management schemes in the evaluation of RhD sensitization.
Authors: Yuelong Guo; Michael P Busch; Mark Seielstad; Stacy Endres-Dighe; Connie M Westhoff; Brendan Keating; Carolyn Hoppe; Aarash Bordbar; Brian Custer; Adam S Butterworth; Tamir Kanias; Alan E Mast; Steve Kleinman; Yontao Lu; Grier P Page Journal: Transfusion Date: 2018-11-20 Impact factor: 3.157