Production and characterization of murine monoclonal antibodies recognizing HLA-DQ polymorphisms obtained by immunizing mice with transfected L cells.

Two polymorphic anti-HLA-DQB1 mAbs, TM 902 and TM 903, have been produced by immunizing F1 mice (Balb/C x C3H) with HLA-DQ-transfected mouse L cells. Cytotoxic analysis on a panel of HLA-typed cell lines has shown that TM 902 reacts with all the DQB1* alleles except DQB1*0501, *0502, and *0503, and DQB1*0601, *0602, *0603, and *0604, whereas TM 903 reacts with the DQB1*0501, *0502, and *0503, DQB1*0601, *0602, *0603, and *0604, and DQB1*0401 and *0402 alleles. The same reactivity pattern has been confirmed by cytofluorimetric analysis. Indirect immunofluorescence with various class-II-transfected cell lines showed no binding of both mAbs to the DR or DP products, suggesting their reactivity to the DQ products. The use of transfectants expressing HLA-DR/DQ heterodimers demonstrates that TM902 and TM903 mAbs are both specific for the DQ-beta chain. Comparison of the amino acid sequences of the DQ-beta chain suggests the involvement of residues 84-90 (QLELRTT) in the formation of TM902 epitope and of residues 54-55 (GR) in the formation of TM903 epitope.     

Characterization and epitope mapping of four HLA class II reactive mouse monoclonal antibodies using transfected L cells and human cells transfected with mutants of DQB1*0302

Abstract: To study epitopes of HLA class II molecules, four mouse monoclonal antibodies (mAbs) 13B6, 17F8, 19A1 and 12G6 were made using HLA-DQ8, DP2 and DP4 expressing mouse transfectants for immunization. Three of the mAbs, 13B6, 17F8 and 19A1, bound to all DQ1, 4, 8 or 9 positive B-lymphoblastoid cell lines (B-LCLs) and transfectants tested, i.e. cells carrying the DQB1 genes 0302-3, 0401-2, 0501-3, 0601-4 and 0609 irrespective of the accompanying DQA1 gene. These DQB1 genes code for the shared amino acids (aa) GVY in position 45–47 of the DQ β chain. DQ1+4+8+9 specific (IIB3) and DQ3 specific (IVD12) reference mAbs inhibited binding of all three mAbs. Testing 13B6, 17F8 and 19A1 with cells made using aa substitutions in various positions of DQP1*0302 indicated involvement of aa 45 in the epitopes of all three mAbs. The last mAb (12G6) bound to all B–LCLs and all DP transfected cells. However, only some DR transfectants and a single DQ transfectant (carrying DQA1 *0201 and DQB1*0202) bound mAb 12G6. This reactivity pattern correlates with a shared sequence of aa (RFDSDVGE) in position 39–46 of DR-and DQ- and 37–44 of DP p chains.     

Immunogenetics of three novel HLA‐Class II alleles: DRB1*03:112, DQB1*03:02:16 and DQB1*03:139

Three novel HLA‐Class II alleles, DRB1*03:112, DQB1*03:02:16 and DQB1*03:139, are described with predicted bearing haplotypes of A*02:01, B*40:01, C*03:04, DRB1*03:112, DQB1*02:01; A*23:01, B*15:01, C*03:03, DRB1*04:01, DQB1*03:02:16 and A*01:01, B*44:02, C*05:01/03, DRB1*04:01, DQB1*03:139. Serological tests showed that the DRB1*03:112 and DQB1*03:139 specificities failed to react as expected with some well‐documented monoclonal antibodies. Subsequent examination of published HLA‐Class II epitopes and inspection of amino acid motifs suggested that epitopes exist that include the positions of their single substitutions (F31C between DRB1*03:01:01:01 and DRB1*03:112, and R48P between DQB1*03:01:01:01 and DQB1*03:139 specificities). This suggests that the reactivity of the monoclonal antibodies used was dependent on these epitopes and that their loss from these rare allele products resulted in their aberrant serology. The new alleles were found after the sequence‐based typing of 32 530 random UK European routine blood donors suggesting that each has a maximum carriage frequency of 0.0031% in the blood donor population resident in Wales.     

昆明彝族人群HLA-DRB1、DQB1基因多态性

目的　调查昆明彝族 HL A- DRB1、DQB1基因的多态性。方法　应用聚合酶链反应 -序列特异性引物基因分型技术 ,对昆明地区 70名彝族健康儿童进行了 HL A- DRB1、DQB1位点的基因分型。结果在 HL A- DRB1位点共检出了 12种等位基因 ,其中等位基因频率大于 10 %的依次为 HL A- DRB1* 12 (33.5 7% )、DRB1* 0 90 1(11.4 3% )、DRB1* 0 4 (11.4 3% ) ,等位基因频率大于 5 %而小于 10 %的依次为 DRB1* 0 1(8.5 7% )、DRB1* 11(7.86 % )、DRB1* 14 (7.14 % )、DRB1* 15 (7.14 % )、DRB1* 0 8(5 % ) ,等位基因频率小于 5 %依次为 HL A- DRB1* 0 3(2 .86 % )、DRB1* 13(2 .14 % )、DRB1* 0 7(1.4 3% )、DRB1* 16 (1.4 3% )。在 HL A- DQB1位点共检出了 7种等位基因 ,其中等位基因频率大于 10 %的依次为 HL A- DQB1*0 30 1(45 % )、DQB1* 0 5 (2 2 .14 % )、DQB1* 0 30 3(12 .14 % ) ,等位基因频率大于 5 %而小于 10 %的依次为DQB1* 0 4 (6 .4 3% )、DQB1* 0 6 (6 .4 3% ) ,等位基因频率小于 5 %的依次为 DQB1* 0 2 0 1(4.2 9% )、DQB1* 0 30 2 (3.5 7% )。结论　昆明彝族 HL A基因多态性分布不同于北方汉族人群 ,也不同于南方汉族人群 ,有其独特性。     

Immunogenetics of heteroclitic recognition of HLA-DQB1 55R eplet specificity by human alloantibody

Heteroclitic antibodies bind to a related antigen with higher affinity than to the immunizing antigen to which they were generated. This uncommon phenomenon is not well characterized for antibodies to HLA antigens. Here we analyzed allosera reactivity from two transplant recipients sensitized to mismatched donor alleles DQB1*06:01 and DQB1*06:02 respectively. Epitope analysis demonstrated the reactivity of both sera was restricted to DQB1*04, 05, and 06 alleles, with a specificity associated with the 55R eplet. Serum from one of these subjects (TE) was significantly more reactive with DQB1*04 alleles than the immunizing DQB1*06:01 or other alleles, a pattern not present in serum from the other patient. Antibody absorption/elution experiments using B cell lines expressing DQB1*06:01 or DQB1*04:02 alleles confirmed that the heteroclitic TE antibody eluted from cells carrying DQB1*06:01 was significantly more reactive with beads carrying the DQB1*04 alleles than with the DQB1*06 or other alleles. The significantly higher reactivity of the heteroclitic alloantibody with DQB1*04 specificity was explained structurally by variations of amino acid residues within 3.5 Å of 55R. These findings have important implications for the interpretation of DQ alloantibody cross-reactivity frequently observed in transplant recipients.     

Suppression by mAbs against DQB1 peptides of in vitro proliferation of AChR-specific T cells from myasthenia gravis patients

It has been indicated that multiple genes, including HLA genes, are collectively involved in the susceptibility to myasthenia gravis (MG). DQB1 alleles represent one of those associated with MG. We have prepared B-cell hybridomas that produce mAbs against peptides corresponding to the tip of the MHC antigen-binding cavity (region 70–90) of alleles DQB1*02, *03, *05 and *06. The mAbs bound to DQ molecules isolated from cells. In the assays using peripheral blood lymphocytes (PBL) from patients with MG, the mAbs against peptides of the correlate HLA DQ sequences inhibited the in vitro proliferation of acetylcholine receptor (AChR)-specific T cells. The results indicate that the function of disease-related MHC alleles may be blocked by directly and selectively targeting the antigen-presenting region on these MHC molecules. The results also suggest that DQ molecules are one of those involved in the restriction of autoimmune anti-AChR responses in MG. The strategy could provide an effective means for immunointervention in MG. It may also potentially be adapted for down-regulation of undesirable immune responses such as in other autoimmune diseases, allergic reactions, or clinical conditions where immune responses to a therapeutic protein develop.     

Suppression by mAbs against DQB1 peptides of in vitro proliferation of AChR-specific T cells from myasthenia gravis patients

It has been indicated that multiple genes, including HLA genes, are collectively involved in the susceptibility to myasthenia gravis (MG). DQB1 alleles represent one of those associated with MG. We have prepared B-cell hybridomas that produce mAbs against peptides corresponding to the tip of the MHC antigen-binding cavity (region 70-90) of alleles DQB1*02, *03, *05 and *06. The mAbs bound to DQ molecules isolated from cells. In the assays using peripheral blood lymphocytes (PBL) from patients with MG, the mAbs against peptides of the correlate HLA DQ sequences inhibited the in vitro proliferation of acetylcholine receptor (AChR)-specific T cells. The results indicate that the function of disease-related MHC alleles may be blocked by directly and selectively targeting the antigen-presenting region on these MHC molecules. The results also suggest that DQ molecules are one of those involved in the restriction of autoimmune anti-AChR responses in MG. The strategy could provide an effective means for immunointervention in MG. It may also potentially be adapted for down-regulation of undesirable immune responses such as in other autoimmune diseases, allergic reactions, or clinical conditions where immune responses to a therapeutic protein develop.     

A novel DRB3 allele (DRB3*0208), a new allelic variant of DRB1*1502 (DRB1*15023) and two new DQB1 (DQBl*03012 and DQB1*0614) alleJes

Abstract: Four novel HLA Class II alleles were identified using CANTYPE reverse hybridization assay. The initial unusual SSO hybridization patterns were confirmed by cloning and sequencing analysis. DRB3*0208 allele is identical to DRB3*0202 except for three nucleotide substitutions (GAT→ AGC) changing codon 57 from Asp to Ser. This polymorphism has so far been undetected in DRB3 alleles. DRB1*15023 differs from DRB1*15021 by a single silent nucleotide substitution (AAC→AAT, both encoding for Asn) at codon 33. This polymorphism has not, until now, been identified in DRB alleles. Compared with DQB1*03011, the novel DQB1*03012 contains a single silent nucleotide substitution (GCA→GCG, both encoding for Ala) at codon 38. Finally, DQB1*0614 allele is identical to DQB1*0603 except for a single nucleotide substitution (TAC→ TTC), changing codon 9 from Tyr to Phe. Polymorphisms observed here in the DQB1*03012 and DQB1*0614 alleles are present in several of the known DQB1 alleles. DRB3*0208, DQB1*03012 and DQB1*0614 may have arisen from gene conversion, but the DRB1*15023 most likely was generated by a point mutation event. DQB1*0614 was detected in three related subjects, while each of the other three new alleles has only been detected once.     

HLA class II alleles and chronic hepatitis C virus infection

The aim of this study was to investigate association of human leucocyte antigens (HLA)-DRB1 and DQB1 polymorphisms with hepatitis C virus (HCV) infection and with the occurrence of severe liver fibrosis/cirrhosis in chronically infected patients. Ninety-nine white patients, from southeast Brazil, with confirmed HCV chronic infection were included in the study. Severe fibrosis/cirrhosis (METAVIR scores F3-F4) was present in 49 patients. HLA-DRB1 specificities and DRB1*11 and DQB1* alleles were determined by PCR-SSP, and their frequencies were compared between patients and a control group of 103 healthy white Brazilian individuals. The results confirmed previous reports of the association of DRB1*11 and DQB1*03 with protection from chronic HCV infection, but did not confirm their association with protection from severe fibrosis/cirrhosis. Furthermore, the results suggested that the polymorphic sites on HLA molecules responsible for protection from chronic HCV infection are encoded not only by the DRB1*1101 and DQB1*0301, as suggested in the literature, but also by other DRB1*11 and DQB1*03 alleles. Thus, we hypothesized that the common polymorphic residues shared by different DRB1*11 and/or DQB1*03 alleles might be responsible for selection of viral epitopes for presentation to CD4(+) T cells, leading to an efficient immune response against the virus.     

Human leukocyte antigen class II allele association to disease progression in Iranian patients with chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is the most frequent type of leukemia in Western countries, but its incidence is low in Asian populations. In the present study we determined the frequency of DRB1 and DQB1 alleles in 87 Iranian CLL patients and 100 healthy controls using a polymerase chain reaction (PCR) technique. An increased frequency of DRB1*07 (p = 0.04), DQB1*06 (p = 0.01) alleles, and DRB1*13/DQB1*03 haplotype (p = 0.01) and decreased frequency of the DQB1*03 (p = 0.01) allele were observed in our patients compared with healthy controls. Comparison between patients with indolent (n = 42) and progressive (n = 38) disease revealed a significant increase in DRB1*04 and DRB5 alleles in progressive patients. Similarly, a higher frequency of DRB5 (p = 0.01) allele was observed in CD38(+) compared with CD38(-) patients. Classification of the patients into immunoglobulin variable region heavy-chain genes mutated and unmutated subtypes did not reveal significant differences for the expression of any of the HLA alleles or haplotypes between these two subtypes. Our findings observed in an Iranian population indicate that CLL could be associated with distinct HLA class II alleles and haplotypes of which the DQB1*06 allele and DRB1*13/DQB1*03 haplotype have not already been reported in CLL patients from other ethnic backgrounds. Some HLA class II alleles may contribute to disease progression in CLL.     

DQB1*0202 and the new DQB1*0203 allele: a fourth pair of DQB1 alleles differing only at codon 57

Amino acid 57 of DQ β chains is of functional importance as it influences peptide binding, is part of B and T cell epitopes, and is associated with susceptibility and resistance to insulin-dependent diabetes mellitus and humoral immunodeficiencies. Polymorphism of codon 57 is conserved in primates and in HLA class II B genes implying that balancing selection operates on this residue. Previously, three DQB1 allele pairs have been described, that only differ at residue 57. In an African-American Black individual with the HLA pheno-type A23, 30; B58, 63; Cw6; DR18, 12; DR52; DQ5, 2, we found a fourth example of this dimorphism; the new DQB1*0203 allele, that was identical to DQB1*0202 except for codon 57, which encodes aspartic acid and alanine respectively in the two alleles. The class II haplotype carrying the new allele was deduced to be DRB1*0302, DRB3*0101, DQA1*05011, DQB1*0203     

寻常型天疱疮的Dsg3特异性抗体反应及其基因限制性

探讨寻常型天疱疮中的自身抗原桥粒芯蛋白（Dsg3)特异性抗体反应及其基因限制性，为自身免疫性疾病机制的研究提供依据。采用RT－PCR法克隆自身抗原Dsg3E1,E2,E3,E4,E5多肽片段的cDNA，定向插入表达载体PGEX－2T，导入大肠杆菌JM109中表达重组融合蛋白并经GST层析柱纯化，进一步经免疫印迹法与PV患者阳性血清反应；应用序列特异性引物聚合酶链式反应（SSP－PCR）技术对HLA－Ⅱ类等位基因进行特异性体外扩增，分析了天疱疮患者HLA基因的DR位点的DRB1、DQB1多态性。Dsg3 E1,E2和E4可与PV患者阳性血清反应，而不与疾病对照组、正常对照组反应。在10个PV患者中，均携带HLA的DR4或／和DR14抗原，有7个DRB1*0402,4个DRB1*1401,7个DQB1*0302,4个DQB1*0503.Dsg3 E1,E2和E4中包含抗体反应相关的抗原表位，HLA DRB1*0402t DQB1＊0302与PV发病中抗E4抗体反应密切相关。     

The HLA DQB 1*0502 allele is neutrally associated with insulin-dependent diabetes mellitus in the Sardinian population

In the Sardinian population a very high incidence of insulin-dependent diabetes mellitus (IDDM) and the lack of HLA-DR2 protective effect due to the high frequency of the A2, Cw7, B17, 3F31, DR2, DQw1 extended haplotype has been reported. This haplotype, carrying a Serine at position 57 of the DQB1*0502 allele, has been previously reported to be underrepresented in patients when compared to controls. In order to provide an explanation for this finding, we defined by RFLP analysis the HLA haplotype of 45 Sardinian IDDM patients and 49 controls. All DR-2DQw1 subjects were molecularly characterized at the HLA DQA and DQB loci. All DR2-positive patients and the vast majority of the DR2-positive controls had the DQB1*0502 allele at the DR2-linked DQB1 locus, with no statistically significant difference between the two groups. All DQA1 genes were the ones expected, with only two exceptions. Nine out of 10 of the DR2-positive patients were compound heterozygotes for DQB1*0201/DQB1*0502 alleles; only this allele combination was significantly increased (p less than 0.0003). Our data suggests that a) the DQB1*0502 allele is neutral for IDDM development and b) the susceptibility to IDDM in our DR2-positive patients is related to the compound heterozygous state between the neutral DQA1*0102/DQB1*0502 and the susceptibility DQA1*0501/DQB1*0201 alleles.     

DNA typing for class II HLA antigens with allele-specific or group-specific amplification. IV. Typing for alleles of the HLA-DR2 group

In this study we present an approach for the definition of the alleles belonging to the HLA-DR2 group by DNA typing with oligonucleotide probes. Following methodology similar to that we used previously for the definition of other HLA-DR subsets, we have now developed primers for DR2-DRB1 and DR2-DRB5 amplification, and probes for the identification of sequences that distinguish the subtypes of this group of genes. The method used defines all the previously described alleles at both DR2-associated DRB loci. In addition, we have identified a variant of DRB1-DR2-Dw2. This new allele has been called DRB1*15.3. It is different from DRB1*1501 in codon 30, where it carries histidine instead of tyrosine. Eight different haplotype combinations of DRB5, DRB1, and DQB1 were identified within the DR2 group and their occurrence in four normal panels of different ethnic origin has been described. Haplotypes containing DRB1*15.3 occurred most frequently in black panel members in whom it was associated with either DQB1*0602 or DQB1*0501. Two unusual haplotypes were observed: one containing elements of DR2-Dw21 (DQB1*0502) and of DR2-Dw22 (DRB1*1602) and one containing elements of Dw21 (DRB1*1601, DQB1*0502) and Dw2 (DRB5*0101). The methods described permit simple and rapid determination of the alleles of the HLA-DR2 group and should be useful for population studies and for investigation of DR2-associated diseases.     

Contribution of a single DQ beta chain residue to multiple sclerosis in French Canadians.

Putative disease susceptibility and resistance HLA class II alleles were studied in 78 French Canadian multiple sclerosis (MS) patients and 79 controls by using sequence-specific oligonucleotide probes to analyze in vitro amplified DNA (PCR-SSOP) typing). In this relatively homogeneous ethnic group, MS was positively associated with DRB5*0101, DQB1*0602, and DQA1*0102 and negatively associated with DQB1*0301. The strongest disease association was with DQB1*0602. Complete DQB1 typing of these individuals, plus RFLP DQ beta typing of an additional five patients showed that 98% of patients compared with 73% of controls carry DQB1 alleles encoding leucine at residue 26. In contrast, 16% of patients compared with 38% of controls carry DQB1 alleles encoding tyrosine at the same residue, and 22% of patients versus 44% of controls carry DQB1 alleles encoding glycine at residue 26. The positive disease correlation was confirmed with SSO probes designed to hybridize to codons for amino acids 22-27 of DQB1*0602, 0603, 0604, 0302, 0303 or to codons for amino acids 25-31 of DQB1*0201; all of these alleles encode Leu 26. These findings suggest that DQ beta chain polymorphisms at a single residue contribute to the development of MS in the French Canadian population.     

HLA-DRB1, -DQB1 and -DPB1 polymorphism in the Slovak population

Abstract: HLA-DRB1, -DQB1 and -DPB1 allele frequencies were investigated in a sample of the Slovak population by PCR-SSP and PCR-RFLP methods. The most frequent DRB1 alleles were DRBl*1101–5 (0.2038), DRBl*0701–2 (0.1423), and DRBl*1501–2 (0.1231). The most rare alleles found were DRBl*0901 (0.0038), and DRB1*1201 (0.015). The most common DQB1 alleles were DQBl*0301 (0.2448), DQB1*0201 (0.2098), and DQB1*0501 (0.1119), respectively. The alleles with the least occurrence rate were DQBl*0601 (0.0035) and DQB1*0401 (0.007). The most common DPB1 alleles found were DPBl*0401 (0.4329), DPBl*0402 (0.2089), and DPB1*0201 (0.1438), respectively. The least frequent alleles were DPBl*0601, *1101, and *1501 (0.0034). Allele frequencies found in our study were compared to those in Czech, Austrian, and German populations. No statistically significant differences were observed.     

Molecular characterization of HLA class II genes in celiac disease patients of Latin American caucasian origin

Abstract: In the present study, the polymorphic domain of HLA class II genes present in a pediatric population of Argentinian celiac disease patients was analyzed by hybridization to sequence-specific oligonucleotides and DNA sequencing. Sixteen out of 16 DR5/7 heterozygous patients bore the DQA1*0501 and DQB1*0201 alleles implicated in the DQ2 risk specificity. The second exon of DQA1, DQB1 and DRB1 genes from 2 DR5/7 patients was characterized by DNA sequencing. The following alleles were found in both patients: DRB1*1101 and DRB1*0701; DQB1*0301 and DQB1*0201; DQA1*0501 and DQA1*0201. Previous serological analysis in this population had shown the presence of DQ2 in 95% of the patients (40% in controls) and a negative association with DQ1 haplotypes, suggesting the presence of other "permissive" or neutral alleles. The following HLA-DQB1 alleles, besides DQB1*0201, were identified in 31 CD patients: DQB1*0301, 0302, 0401 and 0402. All these alleles share a common pattern of residues between positions 84 and 90, and distinct from that present in DQ1-related alleles.     

Susceptible and protective associations of HLA DRB1*/DQB1* alleles and haplotypes with ischaemic stroke

Stroke has emerged as the second commonest cause of mortality worldwide and is a major public health problem. For the first time, we present here the association of human leucocyte antigen (HLA)‐DRB1*/DQB1* alleles and haplotypes with ischaemic stroke in South Indian patients. Ischaemic stroke (IS) cases and controls were genotyped for HLA‐DRB1*/DQB1* alleles by polymerase chain reaction sequence‐specific primers (PCR‐SSP) method. The frequencies of HLA class II alleles such as DRB1*04, DRB1*07, DRB1*11, DRB1*12, DRB1*13, DQB1*02 and DQB1*07 were high in IS patients than in the age‐ and gender‐matched controls, suggesting that the individuals with these alleles are susceptible to ischaemic stroke in South India. The frequencies of alleles such as DRB1*03, DRB1*10, DRB1*14, DQB1*04 and DQB1*05 were less in IS cases than in the controls, suggesting a protective association. Haplotypes DRB1*04‐DQB1*0301, DRB1*07‐DQB1*02, DRB1*07‐DQB1*0301, DRB1*11‐DQB1*0301 and DRB1*13‐DQB1*06 were found to be high in IS patients conferring susceptibility. The frequency of haplotype DRB1*10‐DQB1*05 was high in controls conferring protection. IS‐LVD and gender‐stratified analysis too confirmed these susceptible and protective associations. Thus, HLA‐DRB1*/DQB1* alleles and haplotypes strongly predispose South Indian population to ischaemic stroke. Further studies in different populations with large sample size or the meta‐analysis are needed to explain the exact mechanism of associations of HLA gene(s) with IS.     

Comprehensive typing of DQB1 alleles by PCR-RFLP

Abstract: The protocols represented in this report can resolve all 22 DQB1 alleles. The second exon of DQB1 was subjected to PCR using two group-specific primers to obtain DQB1 group 1 (DQ5 and DQ6) and group 2 (DQ2, DQ3, DQ4) specific amplified products, respectively. Three endo-nucleases, Apal, BssHII and Ncil, can provide typing of DQ5 and DQ6 based on easy-to-read uncleaved, cleaved and a combination of uncleaved/cleaved patterns. Similarly, two endonucleases, FokI and BgII can define the specificities DQ2, DQ3 and DQ4. Moreover, all 13 group 1 DQB1 alleles and all but one of their 78 possible heterozygotes can be unambiguously resolved using an extended panel of 10 endonucleases. The remaining pair of heterozygotes, DQB1*05031/0603 and 05032/0608, can however be resolved by double digestion with BsmFI and SfaNI. RsaI splits the previously unresolved alleles DQB1*0602 and 0603 in the amplified products of the modified primer SDQ-01. Fnu4HI can resolve DQB1*0606 from 0605. DQB1*0603, 0607 and 0608 can be resolved by SfaNI and the new endonuclease BsmFI. The comprehensive typing of group 2 DQB1 alleles can be achieved using five endonucleases. All 9 group 2 DQB1 alleles and all but one pair (DQB1*0301/0302 from DQB1*03032/0304) of 36 possible heterozygotes can be resolved. Thus, PCR-RFLP remains a simple, inexpensive and reliable method for DQB1 typing. The PCR-RFLP can be used for comprehensive DQB1 typing either independently or to complement the PCR-SSP and PCR-SSOP methods.     

Genetic differences in HLA-DQA1* and DQB1* allelic distributions between celiac and control children in Santiago, Chile.

Celiac disease is a permanent gluten intolerance strongly associated with HLA class II antigens. The over presentation of particular HLA alleles and haplotypes has been described in several populations. Different lines of evidence obtained during the last years suggest that a particular HLA-DQ heterodimer, encoded by the DQA1*0501 and DQB1*0201 genes in cis or trans conformation, confers the primary disease susceptibility. We report the HLA class II allelic distribution and DQA1/ DQB1 genotypes in 62 Chilean celiac patients compared with 124 control subjects in Santiago, Chile. We found a pronounced increase of the "susceptible" alleles :DQA1*0501 (0.480 vs 0.169, Pc < 0.0005), DQB1*0302 (0.430 vs 0.242, Pc = 0.002) and DQB1*0201 (0.250 vs 0.125, Pc = 0.037) in celiac patients in comparison with control children. As for "protective" alleles, we detected a high frequency of DQA1*0101 (0.310 vs 0.160, Pc = 0.01), DQA1*0201 (0.105 vs 0.010, Pc < 0.0075) and DQB1*0301 (0.250 vs 0.100, Pc = 0.010) in controls. In relation to risk haplotypes, the main combination observed was the conformation DQ8 (DQB1*0302/DQA1*0301) over DQ2 (DQB1*0201/DQA1*0501). In conclusion, results show that celiac disease in Chilean patients is primarily associated with DQ8 conformation. This is concordant with the high frequency of DR4 alleles (in linkage disequilibrium with DQB1*0302) detected in Amerind groups in Chile, where DQB1*0302 is more frequent than DQB1*0201.