Objective: To evaluate the operational efficiency of a single-center platelet genotype database in HLA genotype-matched platelet search and applcation, and to compare the efficiency among different matching strategies, hospitals, and between first-time versus repeat application modes. Methods: Data on platelet genotype-based matching applications from December 1, 2025, to May 31, 2026, at our center were collected. Four time intervals were defined according to the workflow: the time from clinical hospital application to blood center receipt confirmation (Δt ); the time from blood center confirmation to completion of platelet gene database search (Δt ); the time from completion of database searching to availability of compatible platelet products (Δt ); and the time from clinical hospital application to availability of compatible platelet products (Δt ). Applications were classified as first-time or repeat based on application history, and as speed-priority matching strategy or classical matching strategy based on matching approach, with efficiency differences compared across the groups. Results: A total of 1 051 valid matching applications were included. The median values of Δt , Δt , Δt , and Δt were 6.2 h, 0.1 h, 0.2 h, and 11.5 h, respectively. Overall, 55.9% of genotype-based matches resulted in compatible platelets being available on the same day. The median Δt of the speed-priority strategy was significantly shorter than that of the classical strategy (5.2 h vs 39.6 h, P<0.001). Median Δt differed significantly among hospitals (P<0.001), with the shortest being 4.4 h. The median Δt for repeat applications was 6.9 h, significantly shorter than that for first-time applications (27.7 h, P<0.001). Conclusion: The platelet gene database demonstrates sufficient efficiency in HLA genotype-matched platelet searching and application to meet clinical demands. Optimizing matching strategies and hospital application workflows may shorten the time for obtaining compatible platelets.

[Objective] To resolve the ambiguities of HLA genotyping generated by next generation sequencing (NGS) using nanopore sequencing technology. [Methods] A total of 38 samples with ambiguous HLA genotyping by NGS in our laboratory were collected, and HLA-A, -B, -C, -DRB1, -DRB3/4/5, -DQA1, -DQB1, -DPA1 and -DPB1 loci in these samples were amplified using primers in the same commercial NGS HLA genotyping kit, then subjected to third-generation library construction, and sequenced on the nanopore sequencer. The sequencing data were converted into Fastq files and analyzed by software, and the genotypes of 11 HLA loci were obtained. The ambiguities were counted directly. [Results] The high-resolution genotyping at the second domain of 11 HLA loci of 38 samples using the third generation sequencing (TGS) were consistent with the results of the NGS method at a rate of 100%. The genotypes for the HLA-A, -B, -C, -DRB3, -DRB4, -DQA1 and -DPA1 loci by TGS were all only one result, and the discrimination rate for ambiguities of the HLA-A, -B, -C, and -DQA1 loci (all caused by the difficulty in phasing due to the short NGS read length) was 100%. Among the HLA-DRB1, -DRB5, -DQB1 and -DPB1 loci, the discrimination rate of TGS for the ambiguities caused by non-amplification of exon 1 was 0% and by the short NGS read length was 100%. [Conclusion] Nanopore technology was used to identify the ambiguities of 11 HLA loci in this study, and the ambiguities caused by the short read length disadvantage of the NGS method could be solved effectively and the accuracy of HLA genotyping would be improved.

Objective:To identify the nucleotide sequence of a novel HLA-DRB1*12: 106 allele. Methods:A blood donor who was joined into the database for platelet matching transfusion at the Blood Center of Zhejiang Province in 2023 was selected as the study subject. HLA genotyping was carried out through next-generation sequencing based on AllType NGS 11 locus, AllType FASTPlex NGS reagents, and Sanger sequencing method. The HLA genotype of the donor by Sanger sequencing and next generation sequencing were assigned by using uTYPE 7.3 and TypeStream Visual 3.0 software, respectively. This study was approved by Medical Ethics Committee of the Zhejiang Blood Center (Ethics No. Provincial Blood Center Ethics Review 2022 Research No. 001). Results:A novel HLA-DRB1*12 allele has been identified, and the full coding sequence has been submitted to the GenBank database (No. OR101190), and the length of submitted sequence was 801 bp, which was officially named as HLA-DRB1*12: 106 by the WHO Nomenclature Committee for Factors of the HLA System (submission No. HWS10066755). Compared with the sequence of the highest homology ( HLA-DRB1*12: 01: 01: 01 allele), a single nucleotide change was identified at position 344 T>G in the exon 2 of the HLA-DRB1*12: 106, which has resulted in replacement of Valine by Glycine at residue 86. The HLA genotype of the proband was determined as HLA-A*02: 01, 11: 01; -B*13: 02, 40: 01; -C*01: 02, 03: 03; -DRB1*07: 01, 12: 106; -DRB3*01: 01; -DRB4*01: 03; -DQA1*02: 01, 04: 01; -DQB1*02: 02, 04: 02; -DPA1*01: 03, 01: 03; -DPB1*02: 01: 02G, 04: 01: 01G. Conclusion:A novel HLA-DRB1 allele has been identified in the Chinese population. The mutated amino acid, located in the peptide binding region of the β chain, may affect the binding characteristics of antigen peptides.

Objective:To investigate a case of antibodies against high-frequency erythrocyte antigens and elucidate the genetic mechanism underlying the blood group.Methods:A Lan-negative patient referred to the Zhejiang Blood Center by Quzhou Hospital of Traditional Chinese Medicine in August 2016 was selected as the study subject. A retrospective study was conducted to collect the proband′s clinical data. The proband′s erythrocyte antigens and unexpected serum antibodies were identified using tube saline and microcolumn agglutination anti-human globulin methods. Antibody specificity was determined by treating erythrocytes with 7 enzymes and 2 chemical reducing agents. Genomic DNA was extracted from the proband′s blood sample for whole genome sequencing (WGS) and erythrocyte blood group gene analysis, with validation by Sanger sequencing. Multiple bioinformatics tools were used to analyze the pathogenicity of the variant. The rare blood group and unexpected antibody specificity were comprehensively determined based on the results of serological and genetic testing. This study has been approved by the Zhejiang Provincial Blood Center Medical Ethics Committee(Ethics No.20190201).Results:The proband was a 91-year-old Han Chinese male with prostatitis, cystitis, and malnutrition in conjunct with emaciation. He had a history of multiple erythrocyte transfusions without observable adverse reactions. Prior to the most recent transfusion, major crossmatch agglutination was observed, which prompted antibody identification. Antibodies against high-frequency antigens were detected in the proband′s serum, with enzyme and reducing agent treatments ruling out antibody specificities associated with 17 blood group systems, e. g., MNS, LU, KEL. WGS analysis identified 4 525 SNPs and 1 046 INDEL variants among erythrocyte blood group genes. Further screening revealed that the proband had a rare blood group due to a homozygous rs755723161 variant. This variant in the ABCB6 gene (c.459delC) has led to a frameshifting mutation (p.Trp154GlyfsTer96), resulting in the Lan-negative rare blood group with a high-frequency antigen deficiency and the production of IgG anti-Lan antibodies in the serum. Conclusion:This study has identified anti-Lan alloantibodies in a Lan-negative patient and, for the first time, elucidated the ABCB6 gene variant underlying the Lan-negative rare blood group in the Chinese population.

Objective:The haplotypes of Killer cell immunoglobulin-like receptors (KIR) can be divided into centromeric and telomeric ones. As the terminal gene at the centromeric end, KIR3DP1 plays an important role in stabilizing the haplotype structure. This study aimed to analyze the distribution of KIR3DP1 gene haplotypes among Han Chinese population in Zhejiang in order provide a basis for further analyzing the role of KIR3DP1 in the KIR haplotypes. Methods:A total of 166 unrelated blood donors from Zhejiang were collected (Blood donation period: March 2020 to August 2020), and genotyping was performed by next-generation sequencing based on exon capture. The copy number and allelic frequency of the KIR3DP1 gene and the distribution of centromeric haplotypes were statistically analyzed. This study was approved by the Medical Ethics Committee of Zhejiang Blood Center (Ethics No.: 2023-001). Results:The KIR3DP1 gene was positive for all individuals but with different copy numbers. Among these, 4 cases (2.4%) had only 1 copy, 156 cases (94.0%) had 2 copies, and 6 cases (3.6%) had 3 copies. A total of 10 KIR3DP1 alleles were found in the population, which could be classified into the KIR3DP1*001-L type, KIR3DP1*003-L type, and KIR3DP1 full deletion type. The KIR3DP1*003-L type allele was linked to the Cen- A01 and Cen- B01 types, and the KIR3DP1*001-L type allele and the KIR3DP1 deletion type were only present in the Cen- B02 type haplotype. Conclusion:This study has derived a high-resolution distribution map of the KIR3DP1 gene in the Han population from Zhejiang, and found that the KIR3DP1 alleles showed different linkage with the centromeric haplotypes, which has provided a basis for further studying the role of KIR3DP1 in genetic immunity.

OBJECTIVE: To identify the nucleotide sequence of a novel HLA-DRB1*12:106 allele. METHODS: A blood donor who was joined into the database for platelet matching transfusion at the Blood Center of Zhejiang Province in 2023 was selected as the study subject. HLA genotyping was carried out through next-generation sequencing based on AllType NGS 11 locus, AllType FASTPlex NGS reagents, and Sanger sequencing method. The HLA genotype of the donor by Sanger sequencing and next generation sequencing were assigned by using uTYPE 7.3 and TypeStream Visual 3.0 software, respectively. This study was approved by Medical Ethics Committee of the Zhejiang Blood Center (Ethics No. Provincial Blood Center Ethics Review 2022 Research No. 001). RESULTS: A novel HLA-DRB1*12 allele has been identified, and the full coding sequence has been submitted to the GenBank database (No. OR101190), and the length of submitted sequence was 801 bp, which was officially named as HLA-DRB1*12:106 by the WHO Nomenclature Committee for Factors of the HLA System (submission No. HWS10066755). Compared with the sequence of the highest homology (HLA-DRB1*12:01:01:01 allele), a single nucleotide change was identified at position 344 T>G in the exon 2 of the HLA-DRB1*12:106, which has resulted in replacement of Valine by Glycine at residue 86. The HLA genotype of the proband was determined as HLA-A*02:01, 11:01;-B*13:02, 40:01;-C*01:02, 03:03;-DRB1*07:01, 12:106;-DRB3*01:01;-DRB4*01:03;-DQA1*02:01,04:01;-DQB1*02:02,04:02;-DPA1*01:03,01:03; -- DPB1*02:01:02G,04:01:01G. CONCLUSION A novel HLA-DRB1 allele has been identified in the Chinese population. The mutated amino acid, located in the peptide binding region of the β chain, may affect the binding characteristics of antigen peptides.
Humans ; HLA-DRB1 Chains/genetics* ; Alleles ; Base Sequence ; Genotype ; Male ; High-Throughput Nucleotide Sequencing ; Blood Donors

OBJECTIVE: To investigate a case of antibodies against high-frequency erythrocyte antigens and elucidate the genetic mechanism underlying the blood group. METHODS: A Lan-negative patient referred to the Zhejiang Blood Center by Quzhou Hospital of Traditional Chinese Medicine in August 2016 was selected as the study subject. A retrospective study was conducted to collect the proband's clinical data. The proband's erythrocyte antigens and unexpected serum antibodies were identified using tube saline and microcolumn agglutination anti-human globulin methods. Antibody specificity was determined by treating erythrocytes with 7 enzymes and 2 chemical reducing agents. Genomic DNA was extracted from the proband's blood sample for whole genome sequencing (WGS) and erythrocyte blood group gene analysis, with validation by Sanger sequencing. Multiple bioinformatics tools were used to analyze the pathogenicity of the variant. The rare blood group and unexpected antibody specificity were comprehensively determined based on the results of serological and genetic testing. This study has been approved by the Zhejiang Provincial Blood Center Medical Ethics Committee(Ethics No.20190201). RESULTS: The proband was a 91-year-old Han Chinese male with prostatitis, cystitis, and malnutrition in conjunct with emaciation. He had a history of multiple erythrocyte transfusions without observable adverse reactions. Prior to the most recent transfusion, major crossmatch agglutination was observed, which prompted antibody identification. Antibodies against high-frequency antigens were detected in the proband's serum, with enzyme and reducing agent treatments ruling out antibody specificities associated with 17 blood group systems, e.g., MNS, LU, KEL. WGS analysis identified 4 525 SNPs and 1 046 INDEL variants among erythrocyte blood group genes. Further screening revealed that the proband had a rare blood group due to a homozygous rs755723161 variant. This variant in the ABCB6 gene (c.459delC) has led to a frameshifting mutation (p.Trp154GlyfsTer96), resulting in the Lan-negative rare blood group with a high-frequency antigen deficiency and the production of IgG anti-Lan antibodies in the serum. CONCLUSION This study has identified anti-Lan alloantibodies in a Lan-negative patient and, for the first time, elucidated the ABCB6 gene variant underlying the Lan-negative rare blood group in the Chinese population.
Humans ; Male ; Blood Group Antigens/immunology* ; Aged, 80 and over ; Retrospective Studies ; ATP-Binding Cassette Transporters

OBJECTIVE: The haplotypes of Killer cell immunoglobulin-like receptors (KIR) can be divided into centromeric and telomeric ones. As the terminal gene at the centromeric end, KIR3DP1 plays an important role in stabilizing the haplotype structure. This study aimed to analyze the distribution of KIR3DP1 gene haplotypes among Han Chinese population in Zhejiang in order provide a basis for further analyzing the role of KIR3DP1 in the KIR haplotypes. METHODS: A total of 166 unrelated blood donors from Zhejiang were collected (Blood donation period: March 2020 to August 2020), and genotyping was performed by next-generation sequencing based on exon capture. The copy number and allelic frequency of the KIR3DP1 gene and the distribution of centromeric haplotypes were statistically analyzed. This study was approved by the Medical Ethics Committee of Zhejiang Blood Center (Ethics No.: 2023-001). RESULTS: The KIR3DP1 gene was positive for all individuals but with different copy numbers. Among these, 4 cases (2.4%) had only 1 copy, 156 cases (94.0%) had 2 copies, and 6 cases (3.6%) had 3 copies. A total of 10 KIR3DP1 alleles were found in the population, which could be classified into the KIR3DP1*001-L type, KIR3DP1*003-L type, and KIR3DP1 full deletion type. The KIR3DP1*003 L type allele was linked to the Cen-A01 and Cen-B01 types, and the KIR3DP1*001*L type allele and the KIR3DP1 deletion type were only present in the Cen-B02 type haplotype. CONCLUSION This study has derived a high-resolution distribution map of the KIR3DP1 gene in the Han population from Zhejiang, and found that the KIR3DP1 alleles showed different linkage with the centromeric haplotypes, which has provided a basis for further studying the role of KIR3DP1 in genetic immunity.
Adult ; Female ; Humans ; Male ; Alleles ; China/ethnology* ; Gene Frequency ; Genotype ; Haplotypes/genetics* ; High-Throughput Nucleotide Sequencing/methods* ; East Asian People/genetics* ; Receptors, KIR/genetics*

OBJECTIVE: To analyze the distribution frequency and characteristics of HLA-A and HLA-B loci in patients with immune-mediated Platelet transfusion refractoriness (iPTR) in order to provide data support for investigating HLA gene matching strategies for platelet transfusion and improving transfusion efficacy. METHODS: A total of 532 iPTR patients who applied for gene matched platelet at the Blood Center of Zhejiang Province between January 2020 and June 2024 were selected as the study subjects. Genomic DNA was extracted from peripheral blood samples from the patients, and the HLA-A and HLA-B loci were detected simultaneously using a PCR-sequence specific oligonucleotide probe method (PCR-SSO) and PCR-sequence based typing (PCR-SBT). Statistical methods were used to analyze the distributions of HLA-A and HLA-B antigens and genotypes on the platelet surface of the patients. An analysis of the differences was conducted to compare the results with the Common and Well-Documented (CWD) allele in the Chinese population from the China Marrow Donor Program (CMDP) database. This study was approved by the Medical Ethics Committee of the Blood Center of Zhejiang Province (Ethics No.: Provincial Blood Center Ethics Review 2023Yan No.004). RESULTS: Among the 532 iPTR patients, 19 HLA-A antigens (including 32 HLA-A alleles) and 37 HLA-B antigens (including 64 HLA-B alleles) were detected. The antigens with the highest frequencies were A2, A11, A24, and B46, B60, B58, with the combined distribution frequency of the top three antigens reaching 71.43% and 36.94%, respectively. The most prevalent alleles of the HLA-A and HLA-B loci were A*11:01, A*24:02, A*02:07 and B*46:01, B*40:01, B*58:01. The frequencies of common alleles A*01:01, A*02:07, A*11:02, A*30:01 and B*13:02, B*27:04, B*40:01, B*44:03, B*46:01 showed significant differences (P < 0.05) compared to the normal population in the CWD table (version 2.4) of CMDP. CONCLUSION The HLA-A and HLA-B genes of the iPTR patients showed great divergence, and the distribution frequencies of certain alleles have differed significantly from those of the normal population. This study has provided genetic data for exploring the molecular mechanism underlying iPTR, which is of significant clinical importance for searching HLA gene matched donors.
Humans ; HLA-B Antigens/genetics* ; Alleles ; Female ; HLA-A Antigens/genetics* ; Male ; Platelet Transfusion/adverse effects* ; Gene Frequency ; Middle Aged ; Adult ; Genotype ; Aged ; Adolescent ; China

A 90-year-old female patient with novel coronavirus infection, severe pneumonia, and no history of blood transfusion andtransplantation.The mixed appearance phenomenon appeared in the admission blood group identification, and was sent the sample to our laboratory for difficult blood group identification. In the tube saline method, the patient′s red blood cells were positively reacted with 2 monoclonal anti-A and 5 human anti-A reagents.In the microcolumn gel method, the patient′s red blood cells showed 2 positive and 2 negative reactions with monoclonal anti-A and 5 positive and 1 negative reactions with human anti-A. The patient ′s red blood cells showed negative reaction with peanuts in phytohemagglutinin, and positive reaction with double flower lentils, wild soybeans and a string of purples. The patient ′s red blood cells treated with papain showed negative reaction with all monoclonal anti-A reagents, human anti-A and phytohemagglutinin. The patient ′s ABO gene was sequenced as ABO * B.01/O.01.02, but C1GALT1C1 gene mutation was not founded in the gDNA of the whole blood sample.It is speculated that the exposure of Tn antigen on the patient ′s red blood cells leads to red blood cells polyagglutination, resulting in ABO blood group inconsistency.