Objective: To explore the association between the HLA antibody positivity rate in female blood donors and pregnancy history, number of pregnancies, interval from the last pregnancy to blood donation, and age, to identify associated variables using a univariate generalized additive model (GAM), and to further analyze the predictive role of characteristic variables for HLA antibody positivity using a random forest model. Methods: HLA antibody detection was performed on 391 female blood donors using the Luminex immunomagnetic bead method. The correlation between pregnancy-related factors and HLA antibodies was analyzed using the Chi-square test. Based on R software, a univariate GAM was first constructed to analyze the association types between characteristic variables and the HLA antibody positivity rate, followed by the construction of a random forest model to evaluate the predictive value of the variables. Results: Among the 391 female blood donors without a transfusion history, the overall HLA antibody positivity rate was 26.34%. The positivity rate in donors with a pregnancy history was significantly higher than that in those without (30.09% vs 9.72%, P<0.05), and HLA antibody positivity rate increased linearly with the number of pregnancies (P<0.05). In the univariate GAM, age and number of deliveries exhibited a non-linear association with the HLA antibody positivity rate (the positivity rate increased sharply between 25-35 years of age and stabilized after 3 deliveries). Besides, the interval from the last pregnancy to blood donation showed a linear association with the HLA antibody positivity rate, and the positivity rate decreased as the interval prolonged (P<0.05). In the random forest model, age (mean decrease gini=29.26) and interval from the last pregnancy to blood donation (mean decrease gini=22.02) were core predictive variables: age was more conducive to identifying positive samples, while the interval from the last pregnancy to blood donation was more helpful for excluding negative samples. The number of deliveries (mean decrease accuracy=16.98) made a significant contribution to predicting positive samples, whereas the number of abortions had no impact. The model had an AUC of 0.583 (95% CI: 0.593 8-0.770 2), indicating a certain predictive value. Conclusion: The associated variables identified by the univariate GAM model, including age, interval from the last pregnancy to blood donation, and number of deliveries, provide a basis for key variables in the random forest model. All three variables have predictive value for HLA antibody positivity, which can provide evidence-based support for personalized transfusion management and stratified screening of female blood donors in this region.

Objective To establish the genetic typing detection technique of matrix-assisted laser de-sorption ionization-time or flight mass spectrometry(MALDI-TOF MS),and to apply it to investigate the polymorphism of the human platelet antigen(HPA)-29-35w low-frequency gene in blood donor population from Guangxi area.Methods The RS number of 7 target gene mutation sites in dbSNP and 21 primers were designed by using Assay Design Suite(ADS)of MassARRAY platform online primer design tool.Seven over-expression vectors inserting into HPA-29-35bb mutant sequence were constructed.Twenty-nine blood donors were randomly selected among the blood donors team in Guangxi area.The samples and vectors conducted the iPLEX Pro multiple genotyping analysis and mass spectrometric detection.Meanwhile,the above samples were sequenced and the sequencing results conducted the comparison validation with the mass spectrometric detec-tion results,then the MALDI-TOF MS genotyping detection technology was established.Then adopting this technology conducted the platelet antigen HPA-29-35W genotyping detection and polymorphism analysis in the samples from 588 blood donors in Guangxi area.Results The MALDI-TOF MS HPA genotyping detec-tion results were consistent with the sequencing results.The HPA-29-35w genotyping results showed that theHPA-29-35w genotype of 588 blood donors in Guangxi area was aa homozygote.Conclusion The genotype detection method of MALDI-TOF MS for HPA-29-35w is successfully established and applied to the screening of HPA-29-35w gene among the blood donor population in Guangxi area.

OBJECTIVETo report on a novel human leukocyte antigen (HLA) allele.

METHODSPolymerase chain reaction-sequence based typing was used for routine HLA typing. For one sample, the result of B locus typing showed mismatch of one base with B*46:01:01, B*15:25:01 at locus 384. The group specific sequencing primers, which target at B*46 and B*15, were used to confirm the difference between the novel allele and the highest homologous allele.

RESULTSThe sequencing results showed that the highest homologous allele to the novel allele was B*46:01:01. The two sequences only differed for position 384 within the exon 3 (384G>T), which resulted in a codon change (GGG>GGT), though the amino acid sequence of the novel allele at position 104 was still Glycine (G). Investigation of the family showed that the novel allele was inherited from the father.

CONCLUSIONThe novel HLA-B allele, discovered in ethnic Zhuangs from Guangxi, has been designated as HLA-B *46:01:18 by the World Health Organization (WHO) HLA Nomenclature Committee.

Adult ; Alleles ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; China ; Exons ; Female ; HLA-B Antigens ; genetics ; Humans ; Male ; Molecular Sequence Data ; Young Adult

Objective To investigate the ambiguity results distribution of HLA-A,B and DRB1 gene sequence-base typing in Guangxi population and to propose the way to resolve.Methods HLA-A,B and DRB1 genes of 1 000 donors in the Guangxi branch bank of China'bone marrow bank were genotyped by PCR-SBT,and then the ambiguity results distribution of the three loci was analyzed.The typing ambiguities resultswere resolved by high-resolution polymerase chain reaction-sequence-specific primers(PCR-SSP) and group specific sequencing primer(GSSP) methods,respectively.Results Among 1 000 samples,at least 1 locus in HLA-A,B and DRB1 genes in 96.7％ samples appeared the ambiguity results,in which the proportions of HLA-A,B and DRB1 loci appearing ambiguity results were 65.7 ％,58.8 ％ and 77.2 ％ respectively.For the samples of detected ambiguity results,single using the GSSP method could resolve the ambiguity typing results of 87.37％ HLA-A,93.54％ HLA-B and 60.49％ HLA-DRB1,using high-resolution PCR-SSP could resolve the ambiguity typing results of 12.63 ％ HLA-A,4.76 ％ HLA-B and 15.29 ％ HLA-DRB1,and the rest 1.70 ％ HLA-B and 24.22 ％ HLA-DRB1 ambiguity results were resolved by both GSSP and high-resolution PCR-SSPs method.Conclusion GSSP and high-resolution PCR-SSPs methods have high abilities to solve HLA ambiguity results both locate inside and outside the sequencing region,respectively.GSSP and high-resolution PCR-SSPs methods are supplement for each other,which can effectively resolve the problem of ambiguity results in high resolution HLA typing.

Objective To investigate the allele and specificity distribution situation of HLA-B15 group and HLA-B40 group antigens among Han population in Guangxi area and to explore their possible influence on transplantation donors selection in clinic.Methods The blood samples of 1 644 Han donors in Guangxi region were performed the HLA-B genotyping by PCR-SBT,the frequencies of each allele were calculated by the direct computing method.The antigen specificity of various alleles were analyzed,then the gene frequencies of HLA-B15 and HLA-B40 groups were compared with those from other populations.Results The gene frequency at HLA-B locus in 1 644 Han persons was inconsistent with the Hardy-Weinberg equilibrium(P<0.05).Fourteen alleles in HLA-B15 group were detected out,which belonged to 5 kinds of antigen specificity.In the HLA-B40 group,6 alleles were detected out,which belonged to two kinds of antigen specificity.Conclusion The antigen polymorphism of HLA-B15 and HLA-B40 groups among Han population in Guangxi area is close to that in southern Chinese Han populations,but which still keeps its characteristics of Guangxi area.

OBJECTIVETo explore the molecular basis for a CD36 deficiency individual and distribution of CD36 gene mutation in Guangxi population.

METHODSA female individual was studied. CD36 phenotype was detected by monoclonal antibody immobilization of platelet antigens assay (MAIPA) and flow cytometry (FCM). The coding regions of the CD36 gene were sequenced. A DNA-based polymerase chain reaction-sequence specific primer (PCR-SSP) assay was used to verify the identified mutation. Cell lines expressing the mutant and wild-type CD36[CD36(MT) and CD36(WT)] were established, with the expression of CD36 determined by Western blotting. The distribution of CD36 gene mutation was investigated among 1010 unrelated individuals with the PCR-SSP assay.

RESULTSBoth MAIPA and FCM assays showed that the patient had type II CD36 deficiency. DNA sequencing showed that she has carried a heterozygous mutation T538C (Trp180Arg) in the exon 6 of CD36. Sequencing of cDNA clone confirmed that there was a nucleotide substitution at position 538 (538T>C). Western blotting also confirmed that the CD36 did not express on the CD36(MT) cell line that expressed the 538C mutant, but did express on the CD36(WT) cell line. The novel CD36 mutation T538C was further verified with 100% concordance of genotyping results by DNA-based PCR-SSP assay and 1010 unrelated individuals. No CD36 538C allele was detected among the 1010 individuals.

CONCLUSIONThis study has identified a novel CD36 mutation T538C(Trp180Arg)(GenBank: HM217022.1), and established a genotyping method for the novel sequence-specific primer PCR. The novel mutation is rare in Guangxi and can cause type II CD36 deficiency.

Base Sequence ; Blood Platelet Disorders ; genetics ; Blood Platelets ; cytology ; metabolism ; Blotting, Western ; CD36 Antigens ; genetics ; metabolism ; Cells, Cultured ; DNA Mutational Analysis ; DNA Primers ; genetics ; Exons ; genetics ; Female ; Flow Cytometry ; Fluorescent Antibody Technique ; Genetic Diseases, Inborn ; genetics ; Genotype ; Genotyping Techniques ; methods ; Humans ; Middle Aged ; Monocytes ; cytology ; metabolism ; Mutation, Missense ; Polymerase Chain Reaction ; methods