Genetic polymorphism and antibody screening of RhD negative Chinese Han population in Jiayuguan / 中国输血杂志

【Objective】 To investigate and analyze the polymorphism of RHD gene in RhD-negative population in Jiayuguan using molecular biological technique, so as to accurately identify RhD-negative individuals, and formulate individualized transfusion strategies. 【Methods】 The RhD negative voluntary blood donors and patients (mainly pregnant women) were recruited. After informed consent, history of blood transfusion and pregnancy of them were investigated, and samples were collected for negative D confirmation, gene sequencing as well as antibody screening and identification. 【Results】 Among the 96 samples, 73 cases were RHD gene deletion, 18 RHD*01EL.01(17 RHD1227A homozygous type and 1 RHD1227A heterozygous type), 2 weak RHD*15 type (845G/A), 1 partial D type, i. e. RHD-CE(7) -D heterozygous allele, and 2 RHD*01N.16 variant. Antibody was detected out in 4 cases, among which 2 were positive for anti-D, 1 anti-D plus anti-E, and 1 anti-Dia. 【Conclusion】 The proportion of DEL gene in RhD negative Chinese Han population in Jiayuguan is slightly lower than that in general Chinese Han population. No anti-D or RHD-HDN was observed in DEL type women due to multiple pregnancy or delivery of D positive newborns.

Identification of two novel weak D alleles by sequence-based typing / 中国输血杂志

【Objective】 To study the molecular basis of D variant and explore the molecular genetic mechanism of novel weak D alleles. 【Methods】 Blood samples were screened for D variants by serological method. The nucleotide sequences of coding region were amplified by PCR and sequenced directly, and RHD gene heterozygosity was detected. 【Results】 Weak D phenotype was confirmed by serological test, and two novel alleles were detected by DNA sequencing. The first was novel weak D 1102A allele, 1102G>A mutation in exon 8, resulting in a 368Glu>Arg substitution in two samples. The second was novel weak D 399C allele, carried a 399G>C mutation in exon 3, which led to a 133Lys>Asn substitution. 【Conclusion】 In this study, D variants were detected by sequence-based typing, and two new alleles were identified.

Epitope analysis and molecular study of weak D type 136: A case report / 中国输血杂志

【Objective】 To identify the blood group epitope of a D variant individual and analyze its molecular characteristics. 【Methods】 The saline test and indirect antiglobulin test (IAT) were used to identify the RhD serologically. The anti-human globulin gel card was used for direct antiglobulin test (DAT). RhD epitopes were detected using the epitope detection kit (D-Screen). RhCE antigens were typed using Rh typing Card. The RHD gene zygomorphism was further analyzed by PCR-RFLP. Ten exons of RHD gene were amplified by PCR and analyzed by direct sequencing. 【Results】 DAT test was negative, and the serological results showed weak expression of RhD, which was D variant. The RhD epitope test results showed that the red blood cells of this patient had a weak agglutination with 4 monoclonal anti-D against epD6.4, epD6.1, epD2.1, and epD5.4 (w+ to 2+ ), and reacted negatively with other epitope antibodies. RhCE antigen typing was Ccee; The RHD gene zygomorphism result was D+ /D-, the sequencing of RHD exons revealed that the first exon carried c. 41C>T (p.Pro14Leu) missense mutation, and its genotype was RHD*01W.136/01N.01. 【Conclusion】 This D variant is the first weak D type 136 reported in the Chinese population, and its phenotype is weak partial D.

Serologic strategy in detecting RHD altered alleles in Brazilian blood donors

ABSTRACT Background: We evaluated different technological approaches and anti-D clones to propose the most appropriate serologic strategy in detecting the largest numbers of D variants in blood donors. Methods: We selected 101 samples from Brazilian blood donors with different expressions of D in our donor routine. The tests were performed in immediate spin (IS) with eleven commercially available anti-D reagents in a tube and microplate. The D confirmatory tests for the presence of weak D included the indirect antiglobulin test (IAT) in a tube, gel and solid-phase red blood cell adherence (SPRCA). All DNA samples were extracted from peripheral blood and the D variants were classified using different molecular assays. Results: The RHD variants identified by molecular analysis included weak D types (1, 2, 3, 11 and 38) and partial Ds (DAR1.2, DAR1, DAR3.1, DAU0, DAU2, DAU4, DAU5, DAU6, DMH and DVII). The monoclonal-monoclonal blend RUM-1/MS26 was the best anti-D reagent used in detecting the D antigen in the IS phase in a tube, reacting with 83.2% of the D variants, while the anti-D blend D175 + 415 was the best monoclonal antibody (MoAb) used in a microplate to minimize the need for an IAT, reacting with 83.2% of the D variants. The D confirmatory tests using SPRCA showed a reactivity (3 - 4+) with 100% of the D variant samples tested. Conclusion: Our results show that, even using sensitive methods and MoAbs to ensure the accurate assignment of the D antigen, at least 17% of our donor samples need a confirmatory D test in order to avoid alloimmunization in D-negative patients.

A case study of severe hemolytic disease with the newborn caused by DⅣb type of RhD variant / 中华微生物学和免疫学杂志

Objective To study a case of severe hemolytic disease with the newborn induced by DⅣb type of RhD variant and to investigate its molecular mechanism .Methods Indirect Coombs test was performed to identify RhD blood type and detect antibodies against red blood cells (RBCs).RHD genes were analyzed by polymerase chain reaction-sequence specific primers (PCR-SSP) analysis.All of the 10 exons of RHD gene were sequenced .The Rhesus boxes were further analyzed to identify the homozygosis of RHD genes.Results The mother of the newborn was RhD positive carrying anti-D antibody.PCR-SSP analysis indicated that the RHD exons 7-9 were missing, although the sequences of other RHD exons were consistent with standard sequences .RHD zygosity test showed that the mother was RHD+/RHD-.The newborn was RhD positive with anti-D antibody in serum .The result of the direct antiglobulin test was also positive .The sequence of the RHD exons 1-10 of the newborn were identical with standard sequences .The genotype of the newborn was identified as RHD+/RHD+homozygote .Conclusion The mother bears a DⅣb genotype lac-king RHD exons 7-9 which is significantly different from the newborn .The anti-D antibodies in the mother might induce the severe hemolytic disease in the newborn .