Objective To investigate the serological characteristics and gene mutation mechanism of RhD variant blood donors in Zhengzhou.Methods From January 2023 to December 2023,1 619 RhD-negative blood donors sent to our labora-tory were selected for the study,and RhD negative confirmation test and RhCE phenotype detection were applied by tube method and microcolumn gel indirect antiglobulin test method.RHD gene amplification and Sanger sequencing were used to detect RhD variant sample genotypes.Results A total of 69 cases of RhD variants were detected in the RhD negative con-firmation test,with a proportion of 4.26%(69/1 619).The RhCE phenotypes were ccEe,Ccee,CcEe and CCee.There were 17 genotypes and 15 phenotypes of the D variant.The RHD?weak partial 15 allele was the most frequent(33 cases),with a frequency of 47.83%(33/69),and the main phenotype was the ccEe.This was followed by the RHD?DVI.3 allele in 20 ca-ses with a frequency of 28.99%(20/69)and the predominant phenotype was Ccee.The RHD?weak partial 15/RHD?01EL.01 heterozygote was found in 3 cases with a frequency of 4.35%(3/69),all with the CcEe phenotype.Other rare genotypes were present in 13 cases with a frequency of 18.84%(13/69).Antibody screening was positive in 3 cases with a frequency of 4.35%(3/69).Two cases of female blood donors,both with history of pregnancy and childbirth,were identified as anti-D;one case of male donor was anti-M.Conclusion The RHD?weak partial 15 genotype was the most common among the RhD variants in blood donors in Zhengzhou,followed by the RHD?DVI.3 genotype.It plays an important role in guarantee-ing the safety of blood supply and guiding precision transfusion.

Objective To analyze the application of serological test results in the diagnosis and treatment of anti-M-in-duced hemolytic disease of the fetus and newborn(HDFN),and to explore HDFN prevention strategies.Methods The se-rological test results of 12 cases of HDFN caused by anti-M diagnosed in our laboratory from January 2017 to December 2023 were retrospectively analyzed,including blood group identification of mothers and children,serum total bilirubin/hemoglo-bin/antibody titer test,and three hemolysis tests in newborns.Clinical data of the children and mothers were collected,in-cluding pregnancy history,blood transfusion history,prenatal antibody testing,history of intrauterine blood transfusion and gestational week of delivery,and the prognosis of the children was followed up.Results All 12 cases of fetal neonatal he-molytic disease due to anti-M were RhD+MN phenotype newborn born to RhD+NN mother,with maternal-fetal incompati-blility in MN blood groups.In the ABO blood group system,ABO incompatibility between mother and child accounted for 41.7%(5/12).None of the mothers had a history of blood transfusion,and the median titer of the test at 4℃was 32,and the median titer at 37℃was 4.The mothers of 3 cases had a history of multiple intrauterine blood transfusions,with an inci-dence of 25%(3/12).One case had an abnormal first pregnancy,with an incidence of 8.3%(1/12),and seven cases had an abnormal pregnancy with a miscarriage,with an incidence of abnormal pregnancy and birth history of 58.3%(7/12).There were 6 cases of premature labor,with an incidence of 50%(6/12).The mothers in three cases underwent regular ob-stetric examination and the specificity of the antibodies was determined,accounting for 25%(3/12).Twelve children had free antibodies with a median titer of 6 at 4℃and 2 at 37℃.Two children had anti-M antibodies that were not reactive at 37℃,with a negative rate of 16.7%(2/12).The positive rate of DAT and elution test was respectively 8.3%(1/12)and 16.7%(2/12)in the children.The median minimum hemoglobin value was 75 g/L,and all 12 children received blood transfusions.The median peak total bilirubin value was 157.5 μmol/L,and none of them reached the threshold for blood ex-change.The rate of delayed anemia was16.7%(2/12),the postnatal mortality rate was8.3%(1/12),and 11 children was free of growth and neurodevelopmental delay in prognosis.Conclusion Anti-M can cause severe HDFN,which can also oc-cur in primigravida.The intensity of antibody titer does not correlate with the severity of the disease,and it is prone to cause delayed anemia,which should be monitored regularly according to the serological characteristics of anti-M and clinical symp-toms,and should be treated timely.

Objective:To serologically and genotypically analyze the pedigree of a case with a new allele c.175_176insGA of B el subtype and preliminarily explore the molecular mechanism of weak expression of glycosyltransferase B. Method:In the descriptive study,a 23-year-old male voluntary blood donor and his family members were selected for the study. The ABO and Le blood types of the proband and his family members was identified by the test tube method. The agglutination inhibition test was applied to detect the B and H antigens in saliva, and the Sanger sequencing and PacBio (Pacific Bioscience) third-generation haplotype sequencing were performed on the study subjects to identify genotypes. Finally, Expasy software were applied to amino acid translation of DNA sequences and prediction of protein length after gene alteration. ORF finder was applied to predict alternative start codons as well as open reading frames of mRNA, and protein expression mechanisms were analyzed.Results:The proband and her sister were B el subtype, her mother was AB el subtype, her father was normal O type, and all members of the family were Le(a+b+) phenotype. Sanger sequencing results showed that a new allele of c.175_176insGA was found in exon 4 of the proband, her mother, and her sister. Third-generation haplotype sequencing detected the haplotypes of the family members, which revealed that the proband was ABO*O.01.02/ABO*BEL.NEW (c.175_176insGA), the father was ABO*O.01.02/ABO*O.01.02, the mother was ABO*A1.02/ABO*BEL.NEW (c.175_176insGA), and the sister was ABO*O.01.02/ABO*BEL.NEW (c.175_176insGA). Analysis of the protein expression mechanism indicated that although the new allele of ABO*BEL.NEW was presumed to cause a frameshift mutation and result in a premature stop codon p.Asp59Glu*fs20 in exon 5, encoding an inactive glycosyltransferase, an alternative start codon could be utilized to initiate translation of B el subtype functional glycosyltransferase. Conclusion:Expression of the new allele of B el subtype is associated with the translation of B el subtype glycosyltransferase initiated by alternative start codons.

【Objective】 To analyze and master the serological and genetic characteristics of the samples with CisAB subtype and their genetic background. 【Methods】 From January 2018 to January 2022, blood samples with discrepant ABO blood typing results, from Zhengzhou voluntary blood donors and hospital patients, were subjected to phenotypic classification using micro column gel card and tube method, as well as amplification of exons 6 and 7 in ABO gene using PCR. The pedigrees of individuals with the same CisAB subtype but different serological typing results in the same family were analyzed. 【Results】 11 The forward typing of 12 samples was AB type, and unexpected antibodies against weaker antigens were found in 11 serum samples, including 9 cases with strong antigen A, 2 cases with strong antigen B, and 1 case with consistent forward and reverse typing results. Gene sequencing confirmed that 11 cases were CisAB01 subtype and 1 case was CisAB05 subtype.Among them, 7 cases had the genotype of CisAB01/O and serological phenotype of A2B3; 2 cases had the genotype of CisAB01/B and phenotype of A2B; 2 cases had the genotype of CisAB01/A and serological phenotype of A1Bx and A1B3; 1 case had the phenotype of AxB. In the CisAB01 family, 1 case of CisAB01/O with A2B3 phenotype and 1 case of CisAB01/B with A2B phenotype were detected. In the CisAB05 family, 2 CisAB05/O01 and 1 CisAB05/O02 were detected. 【Conclusion】 The serological phenotypes of different individuals in the same CisAB01 family can be different when paired with different ABO alleles. It is advisable to accurately identify the CisAB subtype genes with molecular biological methods to ensure blood transfusion safety.

【Objective】 To carry out serological and molecular biological identification of B (A) subtype, and discuss the rational blood transfusion strategy. 【Methods】 Serological and direct sequencing methods were used to detect serotype and genotype of 7 cases of B (A) subtype, and cross matching was performed by saline medium and anti human globulin card to analyze the red blood cells(RBCs) transfusion strategy. 【Results】 The serology results of blood type of 7 samples were similar, with B(A)04/O01 in 3 cases, B(A)04/O02 in 2 cases and B(A)02/O01 in 2 cases. 7 cases of B (A) subtypes were matched with randomly selected blood donors of type O and B on the major side. 【Conclusion】 B(A) subtypes should be identified by genotyping techniques. Washed RBCs of type B and O can be used for B(A) blood type transfusion.