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 explore the effects of different methods on antibody detection through investigating the causes of cross-matching incompatible in a patient with gastric malignant tumor, and to establish flow cytometry protocol for confirming hemolytic transfusion reaction (HTR). [Methods] Antibodies in the patient's serum were identified by red blood cells (RBCs) blood grouping, antibody screening and identification, acid elution test and PEG enhancement test. To confirm HTR, patient RBCs, proximal and distal ends RBCs, separated by capillary centrifugation, were tested by direct antiglobulin test (DAT) and Jk^a antigen single label and double label flow cytometry. [Results] Routine serological technology revealed the presence of anti-C, e (titer:2) and anti-Jka (titer >1) in the patient’s serum. After separation using capillary centrifugation technology, both the proximal and distal DAT and Jk^a antigen tests were negative. Both DAT and Jk^a antigen positive red blood cells (0.21%, 6/6 327) were found in the patient's blood samples by flow cytometry. After separation of blood samples by capillary centrifugation, there were significantly more DAT and Jk^a antigen double-positive RBCs in the distal end (0.43%, 33/7 707) than in the proximal end (0.09%, 15/7 225). Two blood samples were screened from over 100 donor blood samples that are compatible with the patient's cross-matching, and the transfusion effect was favorable. [Conclusion] Serological methods combined with flow cytometry could improve the sensitivity of antibody detection, provide a more accurate basis for the diagnosis of HTRs, and guarantee the safety of blood transfusion.