HLA-DRB3 Antibody
Description
Definition and Biological Role of HLA-DRB3
HLA-DRB3 is a gene encoding the beta chain of the HLA-DR heterodimer, which pairs with the HLA-DRA alpha chain to form a functional MHC class II molecule . These molecules are expressed on antigen-presenting cells (APCs) and present extracellular peptides to CD4+ T cells, initiating adaptive immune responses . Key features include:
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Gene Structure: Six exons encoding leader peptides, extracellular domains, transmembrane regions, and cytoplasmic tails .
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Polymorphism: The beta chain determines peptide-binding specificity, with polymorphisms critical for histocompatibility in transplants .
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Expression: DRB3 is expressed at lower levels than DRB1 but is functionally significant in B cells and monocytes .
HLA-DRB3 Antibody Characteristics
HLA-DRB3 antibodies are polyclonal or monoclonal reagents used to detect and study the DRB3 protein. Notable examples include:
| Antibody | Host | Applications | Reactivity | Molecular Weight | Reference |
|---|---|---|---|---|---|
| ABIN2856027 | Rabbit | WB, ELISA | Human | 26–28 kDa | |
| 16789-1-AP | Rabbit | WB, IF/ICC, ELISA | Human | 28–30 kDa |
Key Applications:
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Western Blot (WB): Detects DRB3 in lysates from B-cell lines (e.g., Daudi, Raji) .
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Transplantation Diagnostics: Identifies HLA mismatches to predict graft rejection risks .
Antibody Prevalence in Transplantation
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Kidney Transplants: ~7% of patients develop HLA-DRB3 antibodies post-transplant, linked to epitope mismatches .
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Antibody Patterns: Multiple reactivity profiles suggest DRB3 harbors distinct immunogenic epitopes .
Functional Insights
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T-Cell Epitopes: DRB3 presents unique peptides from pathogens like tetanus toxin and influenza, distinct from DRB1-restricted epitopes .
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Dose-Dependent Effects: The HLA-DRB301:01 allele correlates with anti-HPA-1a antibody levels in maternal-fetal incompatibility, influencing neonatal platelet counts .
Expression and Immunogenicity
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mRNA Levels: DRB3 expression is 2–3.5-fold lower than DRB1 in monocytes and B cells, yet sufficient for antigen presentation .
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Surface Staining: Flow cytometry confirms higher DRB3 expression in B cells compared to monocytes .
Allelic Diversity and Technical Considerations
Over 70 HLA-DRB3 alleles have been identified (e.g., 01:01, 02:02), with implications for antibody specificity . Detection assays like Luminex® use allele-specific beads (e.g., DRB3*01:01, *02:02) to quantify antibody titers (MFI ≥1,000 considered positive) .
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
HLA-DRB3 typically presents extracellular peptide antigens ranging from 10 to 30 amino acids in length, derived from the proteolysis of endocytosed antigens in lysosomes. In the tumor microenvironment, it presents antigenic peptides primarily generated in tumor-resident APCs, likely through phagocytosis of apoptotic tumor cells or macropinocytosis of secreted tumor proteins. It also presents peptides derived from intracellular proteins trapped in autolysosomes after macroautophagy, a mechanism particularly relevant for T cell selection in the thymus and central immune tolerance.
The selection of immunodominant epitopes follows two processing modes: 'bind first, cut/trim later' for pathogen-derived antigenic peptides and 'cut first, bind later' for autoantigens/self-peptides. The anchor residue at position 1 of the peptide N-terminus, usually a large hydrophobic residue, is crucial for high-affinity interaction with MHCII molecules.
**Allele DRB3*01:01:** Exclusively presents several immunogenic epitopes derived from C. tetani neurotoxin tetX, playing a significant role in immune recognition and long-term protection. It presents viral epitopes derived from HHV-6B U11, TRX2/U56, and U85 antigens to polyfunctional CD4-positive T cells with cytotoxic activity implicated in controlling HHV-6B infection.
**Allele DRB3*02:02:** Exclusively presents several immunogenic epitopes derived from C. tetani neurotoxin tetX, playing a significant role in immune recognition and long-term protection. Upon EBV infection, it presents to CD4-positive T cells latent antigen EBNA2 (PRSPTVFYNIPPMPLPPSQL) and lytic antigen BZLF1 (LTAYHVSTAPTGSWF) peptides, driving oligoclonal expansion and selection of virus-specific memory T cell subsets with cytotoxic potential to directly eliminate virus-infected B cells. It also presents viral epitopes derived from HHV-6B U11, gB/U39, and gH/U48 antigens to polyfunctional CD4-positive T cells with cytotoxic activity implicated in controlling HHV-6B infection. It plays a minor role in CD4-positive T cell immune response against Dengue virus by presenting conserved peptides from capsid and non-structural NS3 proteins. It displays peptides derived from IAV matrix protein M, suggesting a role in protection against IAV infection. In the context of tumor immune surveillance, it may present to T-helper 1 cells an immunogenic epitope derived from tumor-associated antigen WT1 (KRYFKLSHLQMHSRKH), likely providing for effective antitumor immunity in a wide range of solid and hematological malignancies. It presents to Vbeta2-positive T-helper 1 cells specifically an immunodominant peptide derived from tumor antigen CTAG1A/NY-ESO-1(PGVLLKEFTVSGNILTIRLTAADHR) and confers protective memory response. In metastatic epithelial tumors, it presents to intratumoral CD4-positive T cells a TP53 neoantigen (HYNYMCNSSCMGSMNRRPILTIITL) carrying G245S hotspot driver mutation and may mediate tumor regression.
**Allele DRB3*03:01:** It presents a series of conserved peptides derived from the M. tuberculosis PPE family of proteins, particularly PPE29 and PPE33, known to be highly immunogenic. It presents immunogenic epitopes derived from C. tetani neurotoxin tetX, playing a role in immune recognition and long-term protection. It displays immunodominant viral peptides from HCV non-structural protein NS2, as part of a broad range T-helper response to resolve infection.
- In contrast to HLA-DRB4*01:01P, the inheritance of HLA-DRB3*01:01 is strongly associated with the propensity for mounting a humoral immune response against fetal HPA-1a antigen. PMID: 28019029
- The first time that the haplotypes A33-DR3 and A33-DR9 were found with an enhanced predisposition to type 1 diabetes in Han Chinese. PMID: 27181214
- HLA-DRB3 affects type 1 diabetes risk and islet autoantibodies. PMID: 26740600
- this mouse model unravels a critical role for HLA-DR3 in generating an autoimmune response to SmD and lupus nephritis in the NZM2328 background. PMID: 26475924
- Haplotyping was done on 91 Southern Europe celiac patients. HLA-DR3-DQ2 without HLA-DR7-DQ2 was present in 62.6%, HLA-DR7-DQ2 without HLA-DR3-DQ2 was present in 16.5% and HLA-DR4-DQ8 without HLA-DQ2 was present in 3.3%. PMID: 24628273
- Children with the HLA haplotype DR3-DQ2, especially homozygotes, were found to be at high risk for celiac disease autoimmunity and celiac disease early in childhood. PMID: 24988556
- HLA-DRB3*02:02 on the DRB1*03:01 haplotype can contribute to type 1 diabetes risk. PMID: 23462545
- Three sets of HLA-DR3-positive haplotypes are designed that provide maximum risk of type 1 diabetes development in Indians and suggest a common Caucasian ancestor from which multiple haplotypes evolve independently. PMID: 23387390
- Heterozygosity of HLA-DRB3*01:01 and HLA-DRB4*01:01 as a potential predictor of fetal neonatal alloimmune thrombocytopenia. PMID: 22671039
- There was a strong sporadic inclusion body myositis association with the carriage of the DRB3*01:01 allele which was accounted for by its linkage disequilibrium with DRB1*03:01. PMID: 23010279
- TNF associations with type 1 diabetes are caused by their linkage disequilibrium with specific HLA-DR3-DQ2 haplotypes in the Indian population. PMID: 22366579
- In addition to confirming one of the top findings in the meta-analysis, TRIM26, RNF5 and HLA-DRB3 have been identified as potential candidate genes for schizophrenia. PMID: 22433715
- There are no significant differences in the HLA-DRB3/B4/B5 homozygosity and heterozygosity rates between Korean males and females in both newborns and adults. PMID: 22531795
- All patients with sporadic inclusion body myositis carried the HLA-DRB1*0301 allele, or its equivalent HLA-DR3 serological specificity PMID: 21507567
- IA2 positivity was associated with HLA-DR4/X and HLA-DR3/4 positivity, and hypothyroidism was linked to HLA-DR4/4. PMID: 20371654
- results suggest that DRB3 plays a significant role in antigen presentation with different epitopic preferences to DRB1. PMID: 19830726
- HLA-DR3 is associated with anti-cyclic citrullinated peptide antibody-negative arthritis and not with anti-CCP-positive arthritis PMID: 16200610
- HLA-DR3 alleles are associated with anti-CCP- disease and with lower levels of anti-CCP antibodies PMID: 16320316
- The HLA-DR locus seems to be associated with the genetic susceptibility to develop SNP in Mexicans. PMID: 16815190
- very pronounced distribution bias (P<10(-5)) of the two major DR3 conserved extended haplotypes, with DR3-B18 predominating in type 1 diabetes and DR3-B8 in celiac disease PMID: 16929349
- German patients were distinct from Italian and North American patients with respect to DQ2 and from the North American patients with respect to B8-DR3-DQ2 HLA in autoimmune hepatitis (AIH) type 1. PMID: 17006990
- HLA-DR and INS VNTR alleles mark both type 1 diabetes disease susceptibility and separate Caucasian parental subpopulations PMID: 17052889
- DR3 is strongly associated with ulcerative colitis protection in a Spanish population. PMID: 17714554
- The researchers found an association between HLA-DR3 and the presence of anti-Ro/La antibodies in patients with SLE. PMID: 17910142
- No particular HLA-DR and DQ alleles, as well as tumor necrosis factor alpha and CTLA-4 genotypes, were associated with CAID. PMID: 18223493
- HLA-DR3 allele may influence susceptibility to the sporadic inclusion body myositis. PMID: 18258695
- Risk of type I diabetes in individuals with extended DR3 haplotype 8.1 is genotype-dependent, possibly associated with DPB1 and HLA class I loci. PMID: 18486765
- T-cell responses associated with neonatal alloimmune thrombocytopenia: isolation of HPA-1a-specific, HLA-DRB3*0101-restricted CD4+ T cells PMID: 19136661
- This study showed that significant expression of HLA-DRB3 genotypes in patients with human high-grade gliomas in Sicily. PMID: 19487887
- interactions between Tg and discrete HLA-DR pocket signatures contribute to the initiation of autoimmune thyroid disease PMID: 19776016
Q&A
What is HLA-DRB3 and what is its biological significance?
HLA-DRB3 is a beta chain component of MHC class II molecules, encoded by genes located on the short arm of chromosome 6 (6p21.3). This protein forms heterodimers with alpha (DRA) chains and is expressed on the surface of antigen-presenting cells (APCs) including B cells, macrophages, and dendritic cells . The primary function of HLA-DRB3 is to present peptide antigens to CD4+ T cells, making it a crucial component of adaptive immune responses . Unlike the ubiquitous HLA-DRB1, the HLA-DRB3 gene is only present in certain HLA haplotypes, meaning individuals may lack the gene entirely or possess one or two copies .
What is the prevalence of HLA-DRB3 in human populations?
The HLA-DRB3 gene is present only in a subset of individuals, contributing to genetic diversity in immune responses . Specific alleles have varying frequencies across populations, with HLA-DRB3*01:01 being one of the most common variants, present in approximately 27.6% of individuals with European ancestry . This variable distribution has significant implications for transplantation compatibility and susceptibility to certain immune-mediated conditions.
How can researchers effectively detect HLA-DRB3 expression?
Detection of HLA-DRB3 expression can be achieved through multiple methodologies:
| Application | Recommended Dilution | Positive Cell Lines |
|---|---|---|
| Western Blot (WB) | 1:5000-1:50000 | Daudi, Raji, Ramos |
| Immunofluorescence (IF/ICC) | 1:200-1:800 | Raji |
| ELISA | Assay-dependent | Human samples |
For optimal results, researchers should titrate the antibody for each specific experimental system . Cell lysis conditions and sample preparation significantly impact the detection sensitivity, particularly when examining naturally expressed levels versus overexpression systems.
How do HLA-DRB3 mismatches impact organ transplantation outcomes?
HLA-DRB3 plays a significant role in transplantation immunology because it is frequently mismatched between donors and recipients. In kidney transplantation cohorts, HLA-DRB3 antibodies have been identified in approximately 7% of patients screened with solid phase assays . These antibodies can develop against mismatched HLA-DRB3 molecules and contribute to rejection processes through multiple immunological pathways. The presence of donor-specific anti-HLA-DRB3 antibodies has been associated with reduced graft survival and increased risk of rejection episodes.
What factors influence antibody formation against HLA-DRB3?
Multiple factors influence antibody formation against HLA-DRB3:
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HLA-DRB1 allele distribution: Certain HLA-DRB1 allele groups in donors show differential frequencies compared to control populations, with some (e.g., HLA-DRB1*11) linked to altered expression levels of HLA-DRB3 .
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Allelic variation in immunogenicity: Different HLA-DRB3 alleles (particularly the HLA-DRB3*01 group) demonstrate variation in immunogenicity independent of expression differences .
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Epitope diversity: Multiple antibody reactivity patterns have been observed in assays, indicating that HLA-DRB3 harbors multiple epitopes that can trigger distinct antibody responses .
These findings suggest that both quantitative (expression levels) and qualitative (specific allelic variants) factors contribute to HLA-DRB3 immunogenicity in transplantation scenarios.
What is the relationship between HLA-DRB3 and HIV-1 transmission?
Research on heterosexual partners where one partner was HIV-1 infected has revealed an intriguing association between HLA-DRB3 discordance and HIV-1 transmission protection. When HLA-DRB3 was present in the HIV donor partner but absent in the recipient partner (immunologic discordance), there was a significant association with lack of HIV-1 transmission . In a study of 45 heterosexual partner pairs, 35% of partner pairs (8/23) where HIV-1 transmission did not occur showed immunological discordance at HLA-DRB3, compared with 0 of 11 partner pairs where HIV-1 transmission did occur (P = .027) .
This phenomenon may be explained by the integration of host HLAs into the HIV-1 envelope, potentially enabling "rejection" of the virus by exposed persons through mechanisms analogous to tissue transplant rejection . These findings suggest potential applications for HLA-DRB3 in HIV vaccine development strategies.
How should researchers design studies to investigate HLA associations with disease transmission?
Based on the HLA-DRB3 and HIV research methodology, effective study designs should include:
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Well-defined exposure groups: Clearly establish exposure status and transmission outcomes.
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Comprehensive HLA typing: Perform high-resolution typing of both partners/subjects.
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Discordance analysis: Examine both presence/absence discordance and allelic differences.
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Statistical power considerations: Account for HLA frequency variations in the study population.
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Multiple HLA loci assessment: Evaluate interactions between different HLA genes and haplotypes.
This methodological approach can be applied to studying other infectious diseases where host genetics may influence transmission dynamics.
How does HLA-DRB3*01:01 impact fetal and neonatal alloimmune thrombocytopenia?
HLA-DRB3*01:01 has been identified as a predisposing factor for human platelet antigen 1a (HPA-1a) immunization, which is responsible for most cases of fetal and neonatal alloimmune thrombocytopenia . Research demonstrates a clear dose-dependent effect of this allele:
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The risk of HPA-1a immunization after delivery of an HPA-1a–positive child is 12.7% in women who are HLA-DRB3*01:01 positive but only 0.5% in women who lack this HLA allele .
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HLA-DRB3*01:01 exhibits a dose-dependent impact on anti-HPA-1a antibody levels in immunized women, with homozygous individuals typically producing higher antibody titers than heterozygous individuals .
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There is an inverse relationship between maternal HLA-DRB3*01:01 dosage and neonatal platelet counts in affected pregnancies .
These findings illustrate how gene dosage of a specific HLA allele can directly influence the severity of an alloimmune condition.
What methodological approaches are used to determine HLA-DRB3*01:01 zygosity?
Determining HLA-DRB3*01:01 zygosity requires a multi-step approach:
| Method | Purpose | Resolution Level |
|---|---|---|
| Sequencing exon 2 of HLA-DRB3 gene | Direct identification of allele | High (second-field) |
| HLA-DRB3 typing | Identification of HLA-DRB3 alleles | First-field |
| HLA-DRB1 typing | Indirect determination of DRB3 presence | First or second-field |
Researchers often use a combination of these approaches, as HLA-DRB3 presence is linked to specific HLA-DRB1 haplotypes . Complete zygosity determination requires evaluating both chromosomes to confirm whether an individual carries zero, one, or two copies of HLA-DRB3*01:01.
How should researchers design antibody experiments to account for HLA-DRB3 allelic diversity?
When designing experiments involving HLA-DRB3 antibodies, researchers should:
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Validate specificity across alleles: Test reactivity with multiple HLA-DRB3 allelic variants, as epitope recognition may vary.
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Consider expression levels: Different cell types and activation states may express varying levels of HLA-DRB3, affecting detection sensitivity .
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Include appropriate controls: Use cell lines with known HLA-DRB3 expression patterns (Daudi, Raji, Ramos cells have been validated for WB detection) .
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Optimize protocols: The recommended dilution ranges (WB: 1:5000-1:50000; IF/ICC: 1:200-1:800) should be titrated for specific experimental systems .
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Account for epitope accessibility: Consider fixation methods that preserve the relevant epitopes, particularly for conformational determinants.
What are the key considerations when interpreting dose-dependent effects of HLA-DRB3*01:01?
When analyzing dose-dependent effects of HLA-DRB3*01:01, researchers should consider:
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Individual variation: Even among homozygous individuals, there can be significant variation in antibody responses. In one study, 4 of 13 women homozygous for HLA-DRB3*01:01 had relatively low anti-HPA-1a levels (1-7 IU/mL) despite the general trend toward higher levels in homozygotes .
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Clinical correlation: Antibody levels correlate with clinical outcomes but with important exceptions. Among homozygous individuals with lower antibody levels, three gave birth to children with normal platelet counts, while one had a child with only mild thrombocytopenia .
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Statistical approach: Global analysis methods should be employed to accurately assess dose effects across heterogeneous populations .
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Interacting factors: Consider other genetic and environmental factors that may modulate the effect of HLA-DRB3*01:01 zygosity.
