ABSTRACT
【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.
ABSTRACT
【Objective】 To explore the characteristics of the D antigen epitope of individuals with RhD variants and the genetic molecular mechanism of gene mutations in Guangzhou. 【Methods】 A total of 59 samples of RhD variants were collected from blood donors and hospitals in Guangzhou from January to August 2019. Serological characteristics of D epitopes were further analyzed using two kinds of monoclonal anti-D reagents and D epitope detection kits, and RHCE phenotypic typing was performed. QuickGene DNA extraction kit was used to extract the genomic DNA of the samples, and PCR-RFLP method was used to analyze the RHD gene zygote type. The RHD gene sequence was detected by multiple ligation-dependent probe amplification(MLPA) genotyping, and the RHD exon(1~10) Sanger sequencing was performed on the samples still in doubt after the above detection. DNAStar/SeqMan analysis software was used for comprehensive analysis. 【Results】 In this group of individuals with RhD variants in Guangzhou, 27.12%(16/59) were detected from blood donors [accounting for 0.007%(16/232 793) of blood donors in Guangzhou during the same period], and difficult samples of patients sent by hospitals for determination accounted for 72.88%(43/59). RHD genotype detection: 40.68%(24/59) were RHD*weak partial 15, 25.42%(15/59) were RHD* DⅥ.3 and 33.90%(20/59) were rare RHD variants [76.92%(10/13) were RhD variants with 2 different alleles]. Serological D-screen revealed a relatively fixed pattern of RHD*DⅥ.3 in anti-D antibody(clone: P3*212 23B10), while the others was negative. The phenotypic distribution of RhD variant CE was Ccee 38.98%(23/59), ccEe 35.59%(21/59), CcEe 25.42%(15/59). 【Conclusion】 Weak partial D15 and DⅥ.3 were the most common RhD variants in Guangzhou Han population, and DⅥ can be preliminarily identified by serological methods such as D-Screen anti-D reagent, while the remaining RhD variants can only be identified by molecular biological methods, and >95% of the RhD variants were C+ or E+ phenotypes.
