title: Molecular RH blood group typing of serologically D-/CE+ donors: the use of a polymerase chain reaction-sequence-specific primer test kit with pooled samples.
abstract: The known presence of RHD blood group alleles in apparently D– individuals who are positive for C or E antigens leads to an appropriate investigation for the RHD gene on the red blood cells (RBCs) of D– blood donors, thus preventing their RBCs from immunizing D– recipients. Ready-to-use polymerase chain reaction–sequence-specific primer (PCR-SSP) typing kits are available and allow single-sample results. The need to perform this testing on a large number of donors affiliated with the Transfusion Department of Udine (Northern Italy) led to the use of molecular genetic RH blood group typing with PCR-SSP test kits and DNA samples mixed in pools. From a population of 35,000 blood donors screened for D antigen by serologic typing, a total of 235 samples, distributed in pools of 5 DNA samples, were investigated. Positive results were reevaluated by opening the pools and retesting single samples. Validation of DNA-pool typing with commercial kits was done. Among 235 genotyped samples, 12 were found to be PCR positive (5.1%), exhibiting DEL genotype and RHD-CE-D hybrid alleles. Our data demonstrate that the use of a PCR-SSP commercial test kit with pooled samples is a helpful and valid method to correctly detect RHD alleles. As a consequence, we reclassified our donors as carriers of potentially immunogenic alleles.
authors.affiliation: Responsible Immunohematology Laboratory, Department of Transfusion Medicine, AOU "S. Maria della Misericordia", Udine, Italy.
authors.forename: Donatella
authors.initials: D
authors.lastname: Londero
authors.forename: Mauro
authors.initials: M
authors.lastname: Fiorino
authors.forename: Valeria
authors.initials: V
authors.lastname: Miotti
authors.forename: Vincenzo
authors.initials: V
authors.lastname: de Angelis
mesh_terms: D001788:Blood Grouping and Crossmatching / Q000379:methods / Q000592:standards; D017931:DNA Primers / Q000235:genetics; D003955:Diagnostic Tests, Routine; D005787:Gene Frequency; D005838:Genotype; D006801:Humans; D007558:Italy; D008403:Mass Screening; D016133:Polymerase Chain Reaction / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D012203:Rh Isoimmunization / Q000097:blood* / Q000453:epidemiology / Q000209:etiology / Q000517:prevention & control; D012204:Rh-Hr Blood-Group System / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D012698:Serologic Tests / Q000592:standards; D065227:Transfusion Reaction
chemical_list: D017931:DNA Primers; D011933:Reagent Kits, Diagnostic; D012204:Rh-Hr Blood-Group System; C026410:Rho(D) antigen