ABSTRACT
BACKGROUND: Antibodies to Gerbich blood group antigens are exceedingly rare and can cause moderate transfusion reactions. Several deletional variants of the GE-gene, that harbors long sequence repeats, enable alloimmunization and formation of naturally occurring antibodies. SUBJECT AND METHODS: A female blood donor and soldier of the German Army without history of pregnancy or transfusion showed an antibody reactive with all test cells except for GE:-2-3 RBC. Thus, anti-Ge2 was suspected. Molecular analysis including fragment length specific PCR, Sanger sequencing and NGS should reveal the molecular background of the deficiency. Segregation of the variant alleles should be demonstrated by family analysis. RESULTS: Compound heterozygosity for GYPC exon 2 (GE*01.-02) and exon 3 (GE*01.-03) deletion was detected in the donor and her sister. The mother had one exon 3 amplicon of reduced length, while the father heterozygously exhibited a truncated GYPC exon 2. NGS clearly demonstrated reduced coverages within the deletional fragments within each family member. The donor and her sister showed the complete absence of a 640 bp fragment. DISCUSSION AND CONCLUSION: Rare GE deletion variants can induce naturally occurring anti-Ge2 in Caucasians. Because of an enhanced risk of injury as soldier autologous RBC of the donor were cryopreserved. The donor and her sibling can give blood for each other because of identical ABO, Rh, and K antigen blood types.
Subject(s)
Anemia, Hemolytic, Autoimmune , Blood Group Antigens , Humans , Pregnancy , Female , Blood Donors , Blood Group Antigens/genetics , Blood Transfusion , Antibodies , PhenotypeABSTRACT
BACKGROUND AND OBJECTIVES: Human neutrophil antigens (HNAs) are categorized into five systems: HNA-1 to HNA-5. Given the importance of neutrophils in immunity, we sought to create awareness of the role of HNA diagnostic services in managing immune neutropenia and transfusion-related acute lung injury. To provide health communities all around the world with access to these services, we conducted a survey to create a directory of these HNA diagnostic services. MATERIALS AND METHODS: An Excel table-based survey was created to capture information on the laboratory's location and was emailed to 55 individuals with known or possible HNA investigation activity. The collected data were then summarized and analysed. RESULTS: Of contacted laboratories, the surveys were returned from 23 (38.2%) laboratories; 17 have already established HNA diagnostic (of them 12 were regular participants of the International Granulocyte Immunobiology Workshop [ISBT-IGIW]), 4 laboratories were in the process of establishing their HNA investigation and the remaining 2 responder laboratories, did not conduct HNA investigations. In established laboratories, investigation for autoimmune neutropenia (infancies and adults) was the most frequently requested, and antibodies against HNA-1a and HNA-1b were the most commonly detected. CONCLUSION: The directory of survey respondents provides a resource for health professionals wanting to access HNA diagnostic services. The present study offers a comprehensive picture of HNA diagnostics (typing and serology), identifying weak points and areas for improvement for the first time. Identifying more laboratories involved in HNA diagnostics with limited access to international societies in the field will globally improve HNA diagnostics.
Subject(s)
Neutropenia , Neutrophils , Adult , Humans , Granulocytes , Antibodies , Surveys and QuestionnairesSubject(s)
Autoimmunity , Neutropenia , Autoimmunity/genetics , Humans , Mutation , Neutropenia/geneticsABSTRACT
Background: Gerbich-negative phenotypes of the Gerbich Blood Group System (ISBT 020) are very rare (with the exception of Papua New Guinea). The Gerbich-negative phenotypes Yus and Gerbich are negative for the antigens Ge2, and Ge2 and Ge3, respectively. In antigen-negative individuals, anti-Ge2 and anti-Ge3 antibodies can be naturally occurring, or are triggered during pregnancies and after transfusions. Previous studies suggested an elevated frequency of Gerbich-negative phenotypes for the Middle East. In the summer of 2015, a large-scale migration of people from the Middle East to Europe occurred raising the issue of question how to guarantee blood supply for patients and manage antenatal care for pregnant women from these countries. Materials and Methods: To investigate the frequency of rare Gerbich-negative phenotypes, 1,665 immigrants to Germany originating from the Middle East were genetically tested for the presence of rare Yus, i.e., GE*01.-02, and Gerbich, i.e., GE*01-03, alleles and compared to results obtained from 507 Germans. Results: Seven Yus GE*01.-02.01 and one Gerbich GE*01.-03.02 alleles were exclusively observed among people from the Middle East, with five of them clustering among 797 Syrians. No such alleles were observed in Germans. A cumulative Yus- and GE*01.-03-type allele frequency of 0.00314 and resultant overall Gerbich-negative phenotype frequency of one among 101,633 Syrians were calculated. Conclusion: This manuscript describes for the first time an exclusively genetic screening for carriers of Gerbich-negative alleles. In conclusion, the Gerbich blood group system should be considered as one causative agent of unusual antibodies to red cell antigens, in routine patients and pregnant women, especially when originating from the Middle East.
ABSTRACT
The new allele differs from DPB1*296:01 by a c.292G>A substitution in exon 2.
Subject(s)
High-Throughput Nucleotide Sequencing , Alleles , Base Sequence , HLA-DP beta-Chains , Humans , Sequence Analysis, DNAABSTRACT
BACKGROUND: CD36 isoantibodies are capable of inducing neonatal alloimmune thrombocytopenia (NAIT) and platelet refractoriness. As to now the CD36 type I deficiency has been reported in East Asian and African individuals. However, it is virtually unknown in Caucasians. The aim of this study was to display the prevalence of the CD36 deficiency within parts of the Arabian population in Germany. Secondly, we are presenting the case of a newborn suffering from NAIT which was induced by CD36 antibody. METHODS: Platelet (p) CD36 was determined by flow cytometry on 1328 samples mainly from individuals of Arabian origin and a family with a neonate affected by NAIT. DNA sequencing was performed on all pCD36-negative samples. RESULTS: Thirty-five (2.64%) of all donor samples were pCD36 negative, 19 (1.43%) had a weak expression. Including only individuals from the Arabian peninsula, frequencies were 3.39% and 1.75%, respectively. CD36 type I deficiency on both platelets and monocytes combined with a CD36 isoantibody were detected in the mother of the NAIT baby. The baby was successfully transfused with two HPA-unselected platelet concentrates. In case of need, two platelet units with a weak pCD36 expression were on hand. A total of 45 different CD36 mutations were detected within pCD36-negative individuals, some being homozygous, most of them only present on one allele. CONCLUSION: The CD36-negative phenotype is present in a significant number of individuals of Arabian origin and enables CD36 isoimmunization in NAIT or refractoriness. Blood transfusion services should be aware of such cases.
Subject(s)
Blood Platelets/pathology , CD36 Antigens/genetics , Thrombocytopenia, Neonatal Alloimmune/genetics , Blood Platelets/metabolism , CD36 Antigens/deficiency , Female , Gene Deletion , Gene Expression , Germany/epidemiology , Humans , Infant, Newborn , Male , Mutation , Pedigree , Polymorphism, Single Nucleotide , Thrombocytopenia, Neonatal Alloimmune/epidemiology , Thrombocytopenia, Neonatal Alloimmune/pathologyABSTRACT
BACKGROUND: The human neutrophil antigen 2 (HNA-2), which is expressed on CD177, is undetectable in 3-5% of the normal population. Exposure of these HNA-2null individuals to HNA-2-positive cells can cause immunization and pro-duction of HNA-2 antibodies, which can induce immune neutropenia and transfusion-related acute lung injury. In HNA-2-positive individuals, neutrophils are divided into a CD177pos. and a CD177neg. subpopulation. The molecular background of HNA-2 deficiency and the bimodal expression pattern, however, are not completely decoded. STUDY DESIGN: An international collaboration was conducted on the genetic analysis of HNA-2-phenotyped blood samples, including HNA-2-deficient individuals, mothers, and the respective children with neonatal immune neutropenia and regular blood donors. RESULTS: From a total of 54 HNA-2null individuals, 43 were homozygous for the CD177 *787A>T substitution. Six carried the CD177 *c.1291G>A single nucleotide polymorphism. All HNA-2-positive samples with >40% CD177pos. neutrophils carried the *787A wild-type allele, whereas a lower rate of CD177pos. neutrophils was preferentially associated with *c.787AT heterozygosity. Interestingly, only the *c.787A allele sequence was detected in complementary DNA (cDNA) sequence analysis carried out on all *c.787AT heterozygous individuals. However, cDNA analysis after sorting of CD177pos. and CD177neg. neutrophil subsets from HNA-2-positive individuals showed identical sequences, which makes regulatory elements within the promoter unlikely to affect CD177 gene transcription in different CD177 neutrophil subsets. CONCLUSION: This comprehensive study clearly demonstrates the impact of single nucleotide polymorphisms on the expression of HNA-2 on the neutrophil surface but challenges the hypothesis of regulatory epigenetic effects being implicated in the bimodal CD177 expression pattern.
ABSTRACT
BACKGROUND AND AIMS: Only little is known about blood groups other than ABO blood groups and Rhesus factors in Arabian countries and Iran. During the last years, increased migration to Central Europe has put a focus on the question how to guarantee blood supply for patients from these countries, particularly because hemoglobinopathies with the need of regular blood support are more frequent in patients from that region. Therefore, blood group allele frequencies should be determined in individuals from Arabian countries and Iran by molecular typing and compared to a German rare donor panel. METHODS: 1,111 samples including 800 individuals from Syria, 147 from Iran, 123 from the Arabian Peninsula, and 41 from Northern African countries were included in a MALDI-TOF MS assay to detect polymorphisms coding for Kk, Fy(a/b), Fynull, Cw, Jk(a/b), Jo(a+/a-), Lu(a/b), Lu(8/14), Ss, Do(a/b), Co(a/b), In(a/b), Js(a/b), Kp(a/b), and variant alleles RHCE*c.697C>G and RHCE *c.733C>G. Yt(a/b), S-s-U-, Velnull, Conull, and RHCE *c.667G>T were tested by PCR-SSP. RESULTS: Of the Arabian donors, 2% were homozygous for the FY *02.01N allele (Fynull), and 15.7% carried the heterozygous mutation. However, 0.8% of the German donors also carried 1 copy of the allele. 3.6% of all and 29.3% of Northern African donors were heterozygous for the RHCE *c.733C>G substitution, 0.4% of the Syrian probands were heterozygous for DO *01/DO *01.-05, a genotype that was lacking in German donors. Whereas the KEL *02.06 allele coding for the Js(a) phenotype was missing in Germans; 0.8% of the Syrian donors carried 1 copy of this allele. 1.8% of the Syrian but only 0.3% of the German donors were negative for YT *01. One donor from Northern Africa homo-zygously carried the GYPB *270+5g>t mutation, inducing the S-s-U+w phenotype, and in 2 German donors a GYPB *c.161G>A exchange, which induces the Mit+ phenotype, caused a GYPB *03 allele dropout in the MALDI assay. The overall failure rate of the Arabian panel was 0.4%. CONCLUSIONS: Some blood group alleles that are largely lacking in Europeans but had been described in African individuals are present in Arabian populations at a somewhat lower frequency. In single cases, it could be challenging to provide immunized Arabian patients with compatible blood.
ABSTRACT
CONTEXT: The structure of the human leucocyte antigen (HLA) peptide-binding clefts strongly contributes to monoglandular and polyglandular autoimmunity (AP). OBJECTIVE: To investigate the impact of amino acid polymorphisms on the peptide-binding interactions within HLA class II and its association with AP. DESIGN: Immunogenetic study. SETTING: Tertiary referral center for autoimmune endocrine diseases. SUBJECTS: 587 subjects with AP, autoimmune thyroid disease (AITD), type 1 diabetes (T1D), and healthy unrelated controls were typed for HLA class II. METHODS: Amino acids within the peptide binding cleft that are encoded by HLA class II exon 2 were listed for all codon positions in all subjects. Overall comparisons between disease and control groups with respect to allele distribution at a given locus were performed by assembling rare alleles applying an exact Freeman Halton contingency table test with Monte-Carlo P values based on 150 000 samples. RESULTS: The Monte Carlo exact Fisher test demonstrated marked differences in all 3 loci, DQA1, DQB1, and DRB1 (P < .0001) between AP and both AITD and controls, as well as between AP type II (Addison's disease as a major endocrine component) and AP type III (T1D + AITD). Differences were also noted between AP and T1D pertaining to the DRB1 allele (P < .041). Seven amino acid positions, DRB1-13, DRB1-26, DRB1-71, DRB1-74, DQA1-47, DQA1-56, and DQB1-57, significantly contributed to AP. Five positions in DQA1 (11, 47, 50, 56, and 69) completely correlated (P < .0001). CONCLUSION: Amino acid polymorphisms within HLA class II exon 2 mediate the AP risk and differentiate between thyroid and polyglandular autoimmunity.
Subject(s)
Amino Acids/genetics , Biomarkers/analysis , Diabetes Mellitus, Type 1/diagnosis , Histocompatibility Antigens Class II/genetics , Polyendocrinopathies, Autoimmune/diagnosis , Polymorphism, Genetic , Thyroiditis, Autoimmune/diagnosis , Case-Control Studies , Diabetes Mellitus, Type 1/genetics , Diabetes Mellitus, Type 1/immunology , Diagnosis, Differential , Female , Follow-Up Studies , Genetic Predisposition to Disease , Humans , Male , Polyendocrinopathies, Autoimmune/genetics , Polyendocrinopathies, Autoimmune/immunology , Prognosis , Thyroiditis, Autoimmune/genetics , Thyroiditis, Autoimmune/immunologyABSTRACT
CONTEXT: The major histocompatibility complex (MHC) strongly contributes to the development of polyglandular autoimmunity (PGA). OBJECTIVE: To evaluate the impact of sex on human leukocyte antigen (HLA) association with PGA for the first time. DESIGN: Cross-sectional immunogenetic study. SETTING: Academic tertiary referral Orphan Disease Center for PGA (ORPHA 282196) and immunogenetics laboratory. SUBJECTS: Patients (158) with coexistent type 1 diabetes and autoimmune thyroid disease (adult type 3 PGA, ORPHA 227982) and 479 unrelated healthy controls. INTERVENTIONS: All 637 white subjects were typed for HLA-A, -B, -DRB1, -DQA1, and -DQB1 alleles at a two-field level. MAIN OUTCOME MEASURES: Modification of the gene-disease association by sex. RESULTS: MHC class I HLA-A association was sex related to both the total white adult type 3 PGA collective (n = 158, P = 0.0065), as well as in PGA patients with autoimmune Hashimoto thyroiditis (n = 91, P = 0.010). Compared with HLA-A*02:01, A*11:01 was over-represented in male patients, yet under-represented in women (OR 1.49, 95% CI 0.55 to 3.88 vs 0.42, 0.12 to 1.17). A*24:02 was under-represented in male but not in female patients (OR 0.37, 95% CI 0.11 to 1.04 vs 1.19, 0.65 to 2.15). With the exclusion of the five most frequent alleles (A*01:01, A*02:01, A*03:01, A*11:01, and A*24:02), the sum of all other identified alleles was under-represented in male patients (OR 0.37, 0.18 to 0.72, P = 0.0046). The strong MHC HLA-B association with PGA (P < 0.0001) was not sex related (P = 0.55). Furthermore, no interaction with sex was observed for the MHC class II HLA-DRB1, -DQA1, and -DQB1 alleles. CONCLUSION: MHC class I HLA-A association with type 3 PGA is significantly affected by sex.
Subject(s)
Biomarkers/analysis , Diabetes Mellitus, Type 1/genetics , HLA-A Antigens/genetics , Histocompatibility Antigens Class I/genetics , Polyendocrinopathies, Autoimmune/genetics , Adult , Case-Control Studies , Cross-Sectional Studies , Diabetes Mellitus, Type 1/immunology , Diabetes Mellitus, Type 1/pathology , Female , Follow-Up Studies , Gene Frequency , Genetic Predisposition to Disease , Haplotypes , Histocompatibility Testing , Humans , Male , Middle Aged , Polyendocrinopathies, Autoimmune/immunology , Polyendocrinopathies, Autoimmune/pathology , Prognosis , Sex FactorsABSTRACT
BACKGROUND AND OBJECTIVE: Antibodies to human neutrophil antigens (HNAs) have been implicated in transfusion-related acute lung injury and allo- and autoimmune neutropenia. To date, five HNA systems are assigned, and during the last decades enormous efforts have been undertaken to identify the underlying genes and to characterize the antigens. This review of the literature will provide the current genetic, molecular and functional information on HNAs. RECENT FINDINGS: New information on alleles and antigens has been added to nearly each of the five HNA systems. HNA-1d has been added as the antithetical epitope to HNA-1c that is located on the glycoprotein encoded by FCGR3B*02 but not by FCGR3B. FCGR3B*04 and *05 now are included as new alleles. A CD177*787A>T substitution was demonstrated as the main reason for the HNA-2-negative phenotype on neutrophils. The target glycoprotein of HNA-3 antibodies could be identified as choline transporter-like protein 2 (CTL2) encoded by SLC44A2. The conformation sensitive epitope discriminates between arginine and glutamine at position 152 resulting in HNA-3a and HNA-3b. An additional Leu151Phe substitution can impair HNA-3a antibody binding. Recently an alloantibody against HNA-4b which discriminates from HNA-4a by an Arg61His exchange of the glycoprotein encoded by the ITGAM gene was reported in neonatal alloimmune neutropenia. An update of the current HNA nomenclature based on the new findings was provided in 2016 by the ISBT Granulocyte Immunobiology Working Party nomenclature subcommittee. CONCLUSIONS: The molecular basis of each of the five HNA antigen systems has been decoded during the past decades. This enables reliable molecular typing strategies, antibody detection and specification as well as development of new assays based on recombinant antigens. However, research on HNA alleles, antigens, and antibodies is not finally terminated and also in the future will add new findings.
ABSTRACT
BACKGROUND AIMS: CD8(+) T cells are part of the adaptive immune system and, as such, are responsible for the elimination of tumor cells. Dendritic cells (DC) are professional antigen-presenting cells (APC) that activate CD8(+) T cells. Effector CD8(+) T cells in turn mediate the active immunotherapeutic response of DC vaccination against the aggressive glioblastoma (GBM). The lack of tumor response assays complicates the assessment of treatment success in GBM patients. METHODS: A novel assay to identify specific cytotoxicity of activated T cells by APC was evaluated. Tumor antigen-pulsed DCs from HLA-A*02-positive GBM patients were cultivated to stimulate autologous cytotoxic T lymphocytes (CTL) over a 12-day culture period. To directly correlate antigen specificity and cytotoxic capacity, intracellular interferon (IFN)-γ fluorescence flow cytometry-based measurements were combined with anti-GBM tumor peptide dextramer staining. IFN-γ response was quantified by real-time polymerase chain reaction (PCR), and selected GBM genes were compared with healthy human brain cDNA by single specific primer PCR characterization. RESULTS: Using CTL of GBM patients stimulated with GBM lysate-pulsed DCs increased IFN-γ messenger RNA levels, and intracellular IFN-γ protein expression was positively correlated with specificity against GBM antigens. Moreover, the GBM peptide-specific CD8(+) T-cell response correlated with specific GBM gene expression. Following DC vaccination, GBM patients showed 10-fold higher tumor-specific signals compared with unvaccinated GBM patients. DISCUSSION: These data indicate that GBM tumor peptide-dextramer staining of CTL in combination with intracellular IFN-γ staining may be a useful tool to acquire information on whether a specific tumor antigen has the potential to induce an immune response in vivo.
Subject(s)
Brain Neoplasms/immunology , Cell- and Tissue-Based Therapy/methods , Dendritic Cells/immunology , Glioblastoma/immunology , Monitoring, Immunologic/methods , Antigens, Neoplasm/immunology , Brain Neoplasms/genetics , Brain Neoplasms/therapy , CD8-Positive T-Lymphocytes/immunology , Female , Glioblastoma/genetics , Glioblastoma/therapy , Humans , Interferon-gamma/metabolism , Lymphocyte Activation , Real-Time Polymerase Chain Reaction , T-Lymphocytes, Cytotoxic/immunologySubject(s)
Amino Acid Substitution , Antigen-Antibody Reactions/genetics , Polymorphism, Single Nucleotide , Rh-Hr Blood-Group System/genetics , Rh-Hr Blood-Group System/immunology , Amino Acid Substitution/genetics , Asparagine/genetics , Blood Donors , Humans , Isoleucine/genetics , Male , Rh-Hr Blood-Group System/metabolism , Serologic TestsABSTRACT
BACKGROUND: The Colton blood group antigens Co(a), Co(b) and Co3 are encoded by the AQP1 gene which produces a water channel forming integral protein. The extremely rare Co-deficiency enables immunisation against the Co3 isoantigen. MATERIALS AND METHODS: Four patients from different regions of Europe who belong to the ethnic minority of Romani (Gypsy) presented with irregular antibodies against a high frequency red blood cell antigen. Positive cross-matches with all red blood cells tested were reported. An Anti-Co3 antibody was identified as the cause of incompatibility in the four cases. The genetic background was determined by polymerase chain reaction typing with sequence-specific primers and by DNA sequencing. RESULTS: The Co(a-b-) phenotype was confirmed in the four patients despite the fact that genotyping revealed the CO*01 allele of the AQP1 gene. A homozygous AQP1 c.601delG mutation, leading to a frame shift and producing a premature stop in the next codon, was responsible for the Co-negative phenotype in all four cases. While one patient was successfully transfused with blood from his sibling with the identical mutation, another case, a baby affected by haemolytic disease of the newborn, recovered without transfusion. DISCUSSION: Despite the difficulties in undertaking a population study to determine the prevalence of this AQP1 c.601delG allele in the ethnic minority of Romani, the observations described in this report clearly suggest an accumulation of this mutation, which causes the Co(a-b-) phenotype, in Romani (Gypsy) patients. Further studies are necessary to prove such an accumulation.
Subject(s)
Aquaporin 1/genetics , Base Sequence , Blood Group Antigens/genetics , Roma/genetics , Sequence Deletion , Adult , DNA Mutational Analysis , Female , Humans , Male , Middle AgedABSTRACT
BACKGROUND: The HNA-3a antigen is an important antibody target in the pathophysiology of transfusion-related acute lung injury (TRALI). It is encoded by the choline transporter-like protein 2 (CTL2) gene, which exists in the two transcript variants TV1 and TV2, differing in the upstream promoter and coding region. Only TV1 has been demonstrated to enable choline transport across the cell membrane. STUDY DESIGN AND METHODS: The aim of this study was to determine the CTL2 transcript pattern in human peripheral blood cells and tissues and its capacity to bind HNA-3a antibodies. RNA was isolated from human whole blood, isolated neutrophils, mononuclear blood cells, leukoreduced platelets, human lung, liver, and colon. After reverse transcription, the single-stranded cDNA was amplified using primer combinations specific for the respective transcript. Plasmids containing the entire CTL2 coding cDNA of the transcript variant TV1 or TV2 served as controls. HEK293T cells expressing both variants were used to determine the binding of HNA-3a antibodies. RESULTS: The shorter TV2 transcript was demonstrated in each RNA sample derived from human peripheral blood tested so far, as well as in human lung and liver, whereas the longer TV1 transcript was only detected in human lung and colon. TV1 and TV2 had the same binding capacity to HNA-3a antibodies. CONCLUSION: The expression of TV1 and TV2 is tissue and cell specific, with peripheral blood cells expressing only TV2. This does not affect binding of HNA-3a antibodies. Whether the unequal expression might be relevant in the pathogenesis of TRALI remains to be investigated.
Subject(s)
Membrane Glycoproteins/genetics , Membrane Transport Proteins/genetics , Acute Lung Injury/etiology , Blood Cells , Cell Line , Humans , Isoantigens/metabolismABSTRACT
BACKGROUND: Neonatal immune neutropenia (NIN) is a rare, but potentially life-threatening, disorder caused by maternal alloantibodies recognizing paternal neutrophil antigens on fetal cells. Alloantibodies directed against the human neutrophil alloantigen system (HNA)-1 located on Fcγ receptor IIIb (FcγRIIIb) are most frequently implicated in NIN. In this report, we describe two cases of NIN with alloantibodies against FcγRIIIb, which did not match one of the known HNA-1a, -1b, or -1c specificities, but define a new antigen, HNA-1d. STUDY DESIGN AND METHODS: Neutrophil-reactive antibodies were detected by agglutination, microscopic immunofluorescence, and monoclonal antibody (MoAb)-specific immobilization of neutrophil antigens (MAIGA) assay. For epitope mapping of FcγRIIIb-reactive antibodies, recombinant chimeric variants of FcγRIIIb were used in the MAIGA assay. Genotyping of FCGR3B was performed by allele-specific polymerase chain reaction. RESULTS: Both mothers were typed FCGR3B*01+, *02-, *03+. Antibody screening revealed the presence of alloantibodies reactive with FcγRIIIb encoded by FCGR3B*02, but not with FcγRIIIb encoded by FCGR3B*03. MAIGA with recombinant, partly chimeric FcγRIIIb variants demonstrated that the antigen recognized by maternal antibodies is characterized by two amino acids, Ala78 and Asp82. Among the FCGR3B alleles, the sequence Ala78---Asn82 is exclusively encoded by FCGR3B *02. CONCLUSION: A previously unrecognized second antigen, HNA-1d, is present on FcγRIIIb encoded by FCGR3B*02. This antigen is characterized by the sequence Ala78---Asn82. It appears that only individuals carrying the HNA-1c phenotype can form anti-HNA-1d alloantibodies. The HNA-1 system now consists of four antigens encoded by three alleles.
Subject(s)
Isoantigens/immunology , Neutropenia/immunology , Neutrophils/immunology , Receptors, IgG/immunology , Adult , Epitope Mapping , Female , GPI-Linked Proteins/genetics , GPI-Linked Proteins/immunology , Genotype , Humans , Infant, Newborn , Isoantibodies/blood , Male , Receptors, IgG/geneticsABSTRACT
BACKGROUND: Antibodies against the human neutrophil alloantigen-3a (HNA-3a) play an important role in transfusion-related acute lung injury. The HNA-3a and -3b alloantigens result from a single-nucleotide exchange in the choline transporter-like protein 2 gene (CTL2). We sought for additional polymorphisms that might impair antibody binding to or genotyping of the HNA-3a or -3b antigens. STUDY DESIGN AND METHODS: CTL2-specific complementary DNA (cDNA) fragments were generated from 67 unrelated blood donors followed by DNA sequencing. Polymerase chain reaction with sequence-specific primers (PCR-SSP) was used to test a higher number of donors for relevant new single-nucleotide polymorphisms (SNPs). The granulocyte agglutination test recommended for HNA-3a antibody detection was performed to check HNA-3a antibody binding to the products of the CTL-2 gene variants. RESULTS: Two new missense mutations were demonstrated in the CTL2 cDNA: a 537C>T* exchange leading to a Leu153Phe amino acid substitution and 988C>T variation predicting Thr301Met change. The inherited 537T variant is located in HNA-3a allele results impaired granulocyte agglutination by four of 14 antibodies tested while 988T remains nearly unaffected. CONCLUSIONS: The Leu153Phe exchange next to the HNA-3a/b defining amino acid position can impede the binding of HNA-3a alloantibodies. The HNA-3a genotyping by PCR-SSP might produce misleading results in HNA-3ab heterozygous individuals with the additional CTL2-537T variation of the HNA-3a antigen. These findings must account for the development of new screening assays.
Subject(s)
Genetic Variation , Isoantigens/genetics , DNA, Complementary/chemistry , Female , Humans , Immune Sera/immunology , Isoantigens/immunology , Male , Polymerase Chain Reaction , Polymorphism, Single Nucleotide , Sequence Analysis, DNAABSTRACT
BACKGROUND: Two plateletpheresis cell separator systems were compared in a paired crossover study with respect to the product quality, the number of platelet (PLT) units per donation, and the donor comfort. STUDY DESIGN AND METHODS: Forty-four female and 47 male donors were distributed to three body weight groups. Double PLT units with 6 x 10(11) PLTs were collected from three Fenwal Amicus Crescendo (AC) and three CaridianBCT Trima Accel (TA) machines. Each donor made one donation on each randomly assigned system and answered a questionnaire on the subjective donor comfort. The answers were scored from 5 (best) to 1 (worst). RESULTS: Based on 182 donations, with 91 donations on AC and TA separators each, 179 runs resulted in double PLT units and three (2xAC, 1xTA) in single units. The white blood cell counts were below 1 x 10(6) in all but eight therapeutic units (8xTA; mean, 1.98 x 10(6)). The mean PLT yield (AC 6.00 x 10(11), TA 5.98 x 10(11)), the collection rate, and the PLT extraction coefficient did not significantly differ between the two devices. Differences of the donor comfort over all groups were only observed for the loudness of the instrument (4.63 AC vs. 4.24 TA, p < 0.001) and the subjective impression of the run time (4.24 AC vs. 4.48 TA, p < 0.05). Male donors greater than 88 kg preferred the TA instruments concerning the impact of the needle, run time, overall experience (p < 0.01 each), and willingness to donate on the same instrument again (p < 0.05). CONCLUSIONS: Only minor differences were observed despite the fact that the AC separators are run with two needles and the TA with one needle.