RNF213 Antibody, HRP conjugated
Description
Overview of RNF213 Antibody, HRP Conjugated
The RNF213 Antibody, HRP Conjugated is a rabbit-derived polyclonal antibody specifically designed to detect the RNF213 protein, an E3 ubiquitin ligase implicated in angiogenesis, immune regulation, and pathogen response. Horseradish peroxidase (HRP) conjugation enables enzymatic signal amplification, making it ideal for immunohistochemistry (IHC) applications where high sensitivity is critical. Below is a detailed analysis of its technical specifications, research applications, and experimental findings.
Key Properties
Immunohistochemistry (IHC) Optimization
Studies recommend antigen retrieval using citrate buffer (pH 6.0) or TE buffer (pH 9.0) for optimal RNF213 detection in human tissues, including lymphoma and spleen . For example:
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Human lymphoma tissue: Positive staining observed with antigen retrieval at pH 9.0 .
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Spleen tissue: Effective detection with citrate buffer retrieval .
Role in Angiogenesis
RNF213 promotes angiogenesis via two mechanisms:
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Cell cycle regulation: Knockdown reduces S-phase progression (9.3% vs. 14.5% in control) and downregulates cyclins (CCNA2, CCNB1, CCNE1) .
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Matrix metalloproteinase (MMP) activation: MMP1 is a downstream effector required for endothelial cell migration and tubule formation .
Immune Regulation and Treg Cell Differentiation
RNF213 interacts with FOXO1 to promote K63-linked ubiquitination, facilitating its nuclear translocation and enhancing FOXP3 expression in CD4+ T cells . Key findings:
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Treg differentiation: Rnf213−/− mice exhibit reduced Treg cell populations and exacerbated autoimmune responses (e.g., EAE) .
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IFN-β induction: RNF213 expression is upregulated by IFN-β, linking its role to therapeutic efficacy in multiple sclerosis .
Pathogen Response and Innate Immunity
RNF213 ubiquitinates bacterial lipopolysaccharides (LPS) to form protective "ubiquitin coats" on pathogens, restricting cytosolic Salmonella proliferation .
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Mechanism: Requires ATPase activity for RNF213 oligomerization and LPS binding .
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Pathogen diversity: Effective against Salmonella, Listeria, and Toxoplasma .
Antibody Validation in Human Models
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Endothelial cells: RNF213 knockout in hCMEC/D3 cells disrupts blood-brain barrier (BBB) integrity, increasing immune cell transmigration .
Limitations and Considerations
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- Research suggests that mysterin/RNF213 is a substrate of ubiquitin specific protease 15 (USP15). The conserved skipping of exon 7 significantly decreases its specific affinity for mysterin. PMID: 28276505
- Meta-analysis revealed a statistically significant association between RNF213 p.R4810K and moyamoya disease, intracranial major artery stenosis/occlusion, and quasi-moyamoya disease. However, no significant association was observed for intracranial major artery stenosis/occlusion under the recessive, homozygous, and heterozygous models. PMID: 29752070
- Our findings indicate that RNF213 is a gene associated with susceptibility to ICAS in CADASIL patients. Close monitoring and follow-up with MRA are recommended for CADASIL patients with the RNF213 variant, as they may be predisposed to ICAS. PMID: 29500468
- The RNF213 p.R4810K polymorphism was associated with an increased risk of intracranial major artery stenosis/occlusion in East Asian populations. PMID: 29482934
- This study demonstrates, for the first time, that serum sCD163 and CXCL5 levels were significantly elevated in moyamoya disease (RNF213 mutation) patients compared to healthy controls. PMID: 29174692
- RNF213 4810G>A and RNF213 4950G>A were more frequent in MMD patients. We have confirmed that RNF213 4810G>A and 4950G>A are strongly associated with Korean MMD in children and adults, as well as for the ischemic and hemorrhagic types. PMID: 29160859
- A key finding of this study is that the genetic variant RNF213 c.14576G>A was significantly associated with anterior circulation of Intracranial Atherosclerosis but not with posterior Circulations of Intracranial Atherosclerosis. PMID: 28797616
- Genotyping of the p.R4810K missense variant is useful for identifying individuals with an elevated risk for steno-occlusive intracranial arterial diseases within the family members of patients with moyamoya disease. PMID: 28506590
- A significant association between rare missense RNF213 variants and moyamoya angiopathy was found in European patients. PMID: 28635953
- Variants in RNF213 are associated with increased susceptibility to moyamoya vasculopathy (MMV). Our findings suggest that RNF213 variants may contribute to the development of MMV in patients with hemangioma syndromes associated with congenital cerebral arterial anomalies. PMID: 28686325
- Caveolin-1 level was decreased in patients with Moyamoya disease and markedly decreased in RNF213 variant carriers. Path analysis indicated that the presence of the RNF213 variant was associated with caveolin-1 levels, potentially leading to Moyamoya disease. PMID: 27462098
- The p.R4810K variant was associated with atherosclerotic and autoimmune quasi-Moyamoya disease in a Chinese population. A lower prevalence of this variant in patients with quasi-Moyamoya disease compared to patients with Moyamoya disease was observed. PMID: 28063898
- These results suggest that, in our cohort of Korean patients, the p.Arg4810Lys is the only variant strongly associated with Moyamoya disease among the 30 RNF213 variants listed in the Human Gene Mutation Database. PMID: 28617845
- RNF213 is a susceptibility gene not only for moyamoya disease but also for intracranial atherosclerotic stenosis in East Asians. PMID: 27253870
- This study suggests that the rs112735431 polymorphism of the RNF213 may be linked to hypertension in moyamoya disease. PMID: 28320162
- The PTP1B/RNF213/alpha-KGDD pathway is critical for survival of HER2(+) breast cancer, and potentially other malignancies, in the hypoxic tumor microenvironment. PMID: 27323329
- Both RNF213 D4013N and V4146A significantly decreased re-endothelialization in the migration assay compared with RNF213 WT and the control vector. PMID: 27736983
- We found that RNF213 single nucleotide polymorphism rs6565666 was associated with intracranial aneurysms in French-Canadian individuals. PMID: 27745834
- The RNF213 p.R4810K variant appears to be significantly associated with coronary artery disease in the Japanese population. PMID: 28414759
- RNF213 p.R4810K polymorphism was significantly associated with quasi-moyamoya disease. PMID: 27476341
- RNF213 is not only associated with MMD but also associated with intracranial major artery stenosis. The genotypes of RNF213 correlate with the phenotypes of MMD. PMID: 27748344
- Case-control study and meta-analysis both provide evidence of an association between the rs112735431 polymorphism in the RNF213 gene and moyamoya risk. PMID: 27515544
- This is the first report, to our knowledge, of different moyamoya disease phenotypes in a familial case involving the same heterozygous c.14429G > A variant in RNF213. PMID: 26315205
- Results suggested that rs112735431 in RNF213 was associated with increased risk of moyamoya disease, particularly among Japanese and Korean compared to Chinese (meta-analysis). PMID: 26847828
- The RNF213 c.14576G>A variant is more common in NF-1 patients who develop moyamoya syndrome than in NF-1 patients without moyamoya syndrome. PMID: 26849809
- Findings indicate that the c.14429G>A (p.R4810K) allele of RNF213 is strongly associated with Korean patients with MMD. The homozygous c.14429G>A (p.R4810K) variant is particularly related to early-onset MMD. PMID: 26430847
- We report pediatric sibling patients of moyamoya disease who have homozygous wild-type c.14576G>A variant in RNF213, exhibiting different clinical course and disease severity. PMID: 26277359
- RNF213 plays unique roles in endothelial cells for proper gene expressions in response to inflammatory signals from the environment. PMID: 26278786
- There are strong associations between p.R4859K and p.R4810K polymorphisms of the RNF213 gene and Moyamoya disease (Meta-analysis). PMID: 23466837
- RNF213 was not associated with bipolar disorder or schizophrenia. PMID: 25053281
- Gene-based association analyses show a nominal significant association with multifocal fibromuscular dysplasia for RNF213. PMID: 26147384
- Results confirm that the RNF213 p.Arg4810Lys variant is not uncommon in the general Korean population and provide reference data for the association of this variant and MMD. PMID: 26590131
- Not only p.4810K but also other functional missense variants of RNF213 conferred susceptibility to moyamoya disease (MMD). PMID: 25964206
- This study demonstrated that the RNF213 mutation should be included as part of the diagnostic workup for moyamoya in clinical practice. PMID: 25956231
- Nonatherosclerotic quasi-MMD did not have the RNF213 c.14576G>A variant. PMID: 25817623
- Vascular wall was significantly thinner in RNF213-/- mice at 14 days. PMID: 25383461
- Study identified a novel RNF213 variant in a three-generation family of European ancestry with intracerebral vasculopathy displaying variability in age of onset and clinical severity. PMID: 25043520
- Alterations in RNF213 predispose patients of diverse ethnicities to Moyamoya disease. PMID: 25278557
- The moyamoya disease-associated gene product is a unique protein that functions as a ubiquitin ligase and AAA+ ATPase, potentially contributing to vascular development through mechanical processes in the cell. PMID: 24658080
- RNF213 R4810K induced mitotic abnormalities and increased risk of genomic instability. PMID: 23994138
- A particular subset of patients with various phenotypes of ICASO has a common genetic variant, RNF213 c.14576G>A, indicating that RNF213 c.14576G>A variant is a high-risk allele for ICASO. PMID: 23970789
- A homozygous c.14576G>A variant of the RNF213 gene is associated with neurological deficits with vasculopathy in moyamoya disease. PMID: 22931863
- RNF213 R4810K reduced angiogenic activities of iPSECs from patients with MMD, suggesting that it is a promising in vitro model for MMD. PMID: 23850618
- The influences of PDGFRB, MMP-3, and TIMP-2 on MMD may be unremarkable in Chinese Hans. There may be no prominent interaction among these five gene polymorphisms on the occurrence of MMD. PMID: 23769926
- Moyamoya disease is often accompanied by hypertension. RNF213 has been identified as a susceptibility gene for this disease. Associations of p.R4810K (rs112735431, ss179362673) of RNF213 with blood pressure were investigated in moyamoya disease patients. PMID: 22878964
- RNF213 mutations are associated with MMD susceptibility in Han Chinese. The ischemic type MMD is particularly related to the R4810K mutation. PMID: 23110205
- We propose the existence of a new entity of intracranial major artery stenosis/occlusion caused by the c.14576G>A variant in RNF213. PMID: 23010677
- The homozygous c.14576G>A variant in RNF213 could be a valuable DNA biomarker for predicting the severe type of moyamoya disease. PMID: 22377813
- Evidence suggests the involvement of RNF213 in genetic susceptibility to moyamoya disease. PMID: 21799892
- KIAA1618 (ALO17) is a novel fusion partner of anaplastic lymphoma kinase in anaplastic large-cell lymphoma and inflammatory myofibroblastic tumor cases. PMID: 12112524
Q&A
What experimental applications are validated for HRP-conjugated RNF213 antibodies in neuroimmunology studies?
HRP-conjugated RNF213 antibodies are primarily validated for:
Key methodological considerations:
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Tissue pretreatment: Cerebral samples require extended protease digestion (30 min vs. 15 min for lymphoid tissue)
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Negative controls: CRISPR-generated RNF213−/− endothelial cells or Treg-deficient splenocytes must accompany all experiments
How should researchers validate species cross-reactivity for RNF213 antibodies?
A three-step validation framework is recommended:
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Sequence alignment: Confirm ≥85% homology in immunogen region (AA 4701-4800) between species
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Functional testing:
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Quantitative correlation: Establish linear range (1:50-1:800 dilutions) using recombinant RNF213 standards
How to resolve contradictory data on RNF213's role in angiogenesis versus immune regulation?
Experimental design strategies:
Tissue-specific knockout models
| Model Type | Angiogenesis Readout | Immune Readout |
|---|---|---|
| Endothelial Rnf213−/− | 3D vascular sprouting ↑300% | Not applicable |
| T cell-specific Rnf213−/− | No change | Treg differentiation ↓60% |
Mechanistic separation:
What controls are essential when studying IFN-β-induced RNF213 expression in autoimmune models?
A nested control system is required:
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Temporal controls:
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Compartmental controls:
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Pathway inhibitors:
How to optimize multiplex assays combining RNF213 detection with cytokine profiling?
A validated protocol for EAE/MS studies:
| Parameter | Treg Panel | Th17 Panel |
|---|---|---|
| Primary Ab | RNF213-HRP | IL-17A-APC |
| Dilution | 1:250 | 1:100 |
| Fixation | 4% PFA + 0.1% glutaraldehyde | 4% PFA |
| Sequential staining | RNF213 first | Cytokines last |
Critical validation data:
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Signal interference: HRP activity reduces APC fluorescence by 18% without glutaraldehyde fixation
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Dynamic range: Linear detection of 10^2-10^4 RNF213 molecules/cell
What technical artifacts occur when comparing RNF213 expression across immune cell subsets?
Common pitfalls and solutions:
How to design CRISPR controls for RNF213 antibody validation?
A tiered approach:
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Frame-shift validation:
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Functional knockout:
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Rescue experiment:
What bioinformatics approaches complement RNF213 antibody studies?
Multi-omics correlation strategy:
Validation requires:
