IL17RB Antibody

What is IL-17RB and what is its significance in biological research?

IL-17RB (Interleukin-17 Receptor B) is a type I membrane protein that functions as a receptor for proinflammatory cytokines, primarily binding to IL-17E (strongly) and IL-17B (weakly). The receptor consists of 502 amino acid residues in humans, with a 17 aa signal peptide, 275 aa extracellular domain, 21 aa transmembrane domain, and 189 aa cytoplasmic tail . IL-17RB is significant in research contexts because:

• It represents one of five members of the IL-17 receptor family

• Its expression is significantly upregulated under inflammatory conditions

• Activation of IL-17RB by its ligands results in nuclear factor kappa-B activation

• It plays critical roles in immune regulation and inflammatory processes

• Recent research indicates its involvement in cancer stem cell (CSC) maintenance and tumorigenesis

Understanding IL-17RB function has implications for research in inflammation, immune disorders, and cancer biology.

How is IL-17RB expression distributed in human tissues?

IL-17RB demonstrates tissue-specific expression patterns that are important to consider when designing experiments:

• High expression: Kidneys and liver

• Moderate to low expression: Testes, brain, small intestine

• Also found in various endocrine tissues

• Expression significantly increases under inflammatory conditions

When conducting tissue-specific research, it's important to note that baseline expression levels vary considerably between tissue types, which can affect experimental design and interpretation of results when using IL-17RB antibodies.

What are the common applications for IL-17RB antibodies in research?

IL-17RB antibodies are versatile reagents that can be used in multiple experimental applications:

When designing experiments, researchers should validate the antibody in their specific experimental system, as optimal dilutions may vary based on sample type, fixation method, and detection system.

What is the molecular weight of IL-17RB in Western blot applications?

When working with IL-17RB in Western blot applications, researchers should expect:

• Calculated molecular weight: 56 kDa

• Observed molecular weight range: 56-60 kDa

• Two isoforms: 56 kDa (isoform 1) and 32 kDa (isoform 2)

If your Western blot shows bands at unexpected molecular weights, consider potential post-translational modifications, alternative splicing variants, or degradation products. The discrepancy between calculated and observed molecular weights is common for many proteins due to post-translational modifications like glycosylation or phosphorylation.

How does IL-17RB expression differ between cancer stem cells and differentiated cells?

Research comparing IL-17RB expression between cancer stem cells (CSCs) and differentiated cells has revealed significant differences:

• IL-17RB expression is approximately 10-fold higher in spheroid cells (enriched for CSCs) compared to adherent naturally growing cells

• IL-17RB expression decreases during forced differentiation of spheroid cells, correlating with decreases in stemness markers (OCT4, NANOG, SOX2, LGR5)

• In magnetic bead sorting experiments, CD133+ cells (CSC marker-positive) show significantly higher IL-17RB expression than CD133- cells

• Flow cytometry analysis confirms co-expression of IL-17RB with CSC markers like Lgr5 and CD133

These findings suggest IL-17RB may serve as a novel CSC marker. When designing experiments to study CSCs, consider using IL-17RB antibodies in conjunction with established CSC markers for more comprehensive characterization.

What signaling mechanisms are activated downstream of IL-17RB?

IL-17RB activates complex signaling cascades that researchers should consider when studying this receptor:

• Binding of IL-17B or IL-17E to IL-17RB activates nuclear factor kappa-B signaling

• IL-17RB interacts with TRAF6 through a specific binding domain

• This interaction promotes TRAF6-Beclin-1 binding

• The IL-17RB-TRAF6 interaction leads to K63-mediated ubiquitination of Beclin-1

• Deletion of the TRAF6-binding domain of IL-17RB abolishes the binding between Beclin-1 and TRAF6

When studying IL-17RB signaling, consider using co-immunoprecipitation experiments to detect protein-protein interactions, and ubiquitination assays to monitor post-translational modifications in the signaling cascade.

How might seemingly contradictory findings about IL-17 signaling in disease states be reconciled?

The IL-17 family shows complex and sometimes contradictory roles in diseases like inflammatory bowel disease (IBD). To address these contradictions:

• Consider different IL-17 family members separately (IL-17A vs IL-17B vs IL-17E)

• Examine cell-type specific responses to IL-17 family cytokines

• Investigate temporal aspects of IL-17 signaling during disease progression

• Use Mendelian randomization (MR) approaches with protein quantitative trait loci (pQTL) to examine causal relationships

• Apply multivariable MR (MVMR) to control for potential confounders

These approaches can help resolve apparent contradictions by providing a more nuanced understanding of how different IL-17 family members contribute to disease pathogenesis through distinct mechanisms.

What are the optimal antigen retrieval methods for IL-17RB immunohistochemistry?

For successful IL-17RB immunohistochemistry, proper antigen retrieval is crucial:

• Primary recommended method: TE buffer pH 9.0

• Alternative method: Citrate buffer pH 6.0

Optimization is essential as different tissues may require adjusted protocols. If signal intensity is suboptimal, consider:

• Extending retrieval time

• Adjusting retrieval temperature

• Testing both recommended buffers with your specific tissue samples

• Including positive control tissues (kidney, breast cancer, or colon tissues)

How can researchers validate specificity of IL-17RB antibodies in their experimental systems?

Validating antibody specificity is critical for meaningful results. For IL-17RB antibodies, consider:

• Positive controls: Use tissues/cells known to express IL-17RB (kidney, testis, K-562 cells)

• Negative controls: Include secondary antibody-only controls

• Peptide competition assays: Pre-incubate antibody with immunizing peptide

• Knockdown validation: Compare staining in IL-17RB knockdown vs. control cells

• Multiple antibody validation: Use antibodies targeting different epitopes

• Multiple detection methods: Confirm findings using different techniques (WB, IHC, IF)

Remember that antibody performance can vary between applications and sample preparation methods, making validation in your specific experimental system essential.

What considerations are important when designing in vivo experiments to study IL-17B/IL-17RB signaling?

In vivo studies of IL-17B/IL-17RB signaling require careful experimental design:

• Animal models: BALB/c nude mice (4-6 weeks) have been successfully used

• Cell preparation: Pre-treat cells with recombinant IL-17B (rIL-17B) at defined concentrations

• Cell numbers: Consider 10², 10³, 10⁴, and 10⁵ cells for tumor-initiating capacity analysis

• Controls: Include PBS-treated controls alongside rIL-17B treatment

• Duration: Monitor until significant tumor development (~20% weight decrease)

• Measurements: Track tumor formation time, volume (V = 0.5 × a × b², where a is longer diameter and b is shorter diameter), and weight

• Analysis: Perform histological and immunohistochemical analysis for proliferation markers (Ki67, cyclin-D1) and stemness markers (Sox9, Sox2)

For statistical robustness, include sufficient biological replicates (n=8 per cell concentration recommended) and randomize animal allocation to groups.

How can researchers quantitatively assess the impact of IL-17B on IL-17RB expression?

To quantitatively evaluate how IL-17B affects IL-17RB expression:

• mRNA quantification:

  • Use qRT-PCR to measure IL-17RB transcript levels before and after IL-17B treatment

  • Normalize to appropriate housekeeping genes

• Protein quantification:

  • Western blot with densitometry analysis

  • Flow cytometry to measure surface receptor density

  • Immunofluorescence with quantitative image analysis

• Functional assays:

  • Measure downstream signaling activation (NF-κB reporter assays)

  • Assess TRAF6-Beclin-1 interaction through co-immunoprecipitation

  • Quantify K63-mediated ubiquitination of Beclin-1

Research has shown that exogenous IL-17B enhances IL-17RB expression, creating a positive feedback loop. Serum IL-17B levels in gastric cancer patients positively correlate with IL-17RB mRNA expression in gastric cancer tissues .

How should researchers interpret multiple bands in Western blots using IL-17RB antibodies?

When multiple bands appear in Western blots:

• Expected bands: 56 kDa (isoform 1) and 32 kDa (isoform 2)

• Potential alternative splicing: A secreted variant of IL-17RB has been identified

• Degradation products: Consider adding additional protease inhibitors

• Post-translational modifications: Phosphorylation or glycosylation can alter migration

• Non-specific binding: Optimize blocking conditions and antibody dilutions

If unexpected bands persist, validate specificity using knockdown or knockout controls and consider using alternative antibody clones targeting different epitopes.

What factors might affect IL-17RB antibody performance in different experimental contexts?

Several factors can influence IL-17RB antibody performance:

• Fixation methods: Formalin fixation may mask epitopes differently than alcohol-based fixatives

• Sample preparation: Fresh vs. frozen tissue processing affects antigen preservation

• Antibody format: Consider whether conjugated antibodies are needed for specific applications

• Blocking reagents: BSA vs. serum can affect background and specificity

• Detection systems: Sensitivity varies between chromogenic, fluorescent, and chemiluminescent methods

• Cell/tissue type: Endogenous expression levels and potential interfering substances vary

For optimal results, systematically optimize each experimental variable and include appropriate controls to validate findings across different experimental conditions.

What emerging research directions are important for IL-17RB antibody applications?

The field of IL-17RB research is evolving rapidly, with several promising directions:

• Cancer stem cell biology: Using IL-17RB antibodies to identify and isolate CSCs

• Therapeutic targeting: Developing antibodies that block IL-17B/IL-17RB signaling

• Biomarker applications: Exploring IL-17RB as a diagnostic or prognostic marker

• Single-cell analysis: Integrating IL-17RB antibodies into single-cell profiling platforms

• Structural studies: Understanding the epitope-paratope interactions for improved antibody design

Researchers should stay current with literature as new applications and methodologies continue to emerge in this dynamic field.

How might contradictory findings about IL-17 signaling in inflammatory diseases be further investigated?

To better understand contradictory findings regarding IL-17 signaling in inflammatory diseases like IBD:

• Separate analysis of IL-17 family members (IL-17A, B, C, E, F)

• Tissue-specific and cell-type-specific knockouts

• Temporal regulation studies across disease progression

• Receptor-specific analysis (IL-17RA, IL-17RB, etc.)

• Integration of genomic data using Mendelian randomization approaches

• Multivariable analyses to control for confounding factors