CCR10 Antibody

What is CCR10 and what cellular functions does it regulate?

CCR10, also known as GPR2, is a G-protein coupled receptor that functions as a receptor for the chemokines CCL27 (SCYA27) and CCL28 (SCYA28). It plays critical roles in directing lymphocyte trafficking to specific tissue sites. CCR10-CCL27 interactions are primarily involved in T cell-mediated skin inflammation processes, while CCR10-CCL28 interactions contribute significantly to the accumulation of IgA antibody-secreting cells (ASCs) on mucosal surfaces . The receptor consists of 362 amino acids with a calculated molecular weight of approximately 38 kDa, though the observed molecular weight typically ranges between 38-42 kDa in experimental contexts .

What are the primary research applications for CCR10 antibodies?

CCR10 antibodies are employed across several experimental techniques to investigate chemokine receptor expression and function. Based on validated applications, CCR10 antibodies are predominantly used in:

• Western Blot (WB) analysis with recommended dilutions of 1:500-1:2000

• Immunohistochemistry (IHC) with recommended dilutions of 1:20-1:200

• Flow Cytometry (FC) as demonstrated in published research

• ELISA applications

It is important to note that optimal dilutions can vary significantly between experimental systems, and researchers should conduct titration experiments within their specific testing paradigms to achieve optimal results.

What tissue samples show reliable CCR10 detection?

Research-validated CCR10 antibodies have demonstrated consistent reactivity with human, mouse, and rat samples. Specifically:

For IHC applications, antigen retrieval using TE buffer (pH 9.0) is recommended, though citrate buffer (pH 6.0) may serve as an alternative method .

What are the optimal protocols for CCR10 detection in tissue samples?

For reliable CCR10 detection in tissue samples via Western blot, researchers should:

• Extract proteins from target tissues (brain tissue from mouse/rat models has demonstrated consistent results)

• Separate proteins via SDS-PAGE

• Transfer to appropriate membrane

• Block with standard blocking buffer

• Incubate with anti-CCR10 antibody at 1:500-1:2000 dilution

• Wash and incubate with appropriate secondary antibody

• Develop using standard detection methods

For immunohistochemistry applications:

• Prepare tissue sections (human spleen tissue has shown reliable results)

• Perform antigen retrieval using TE buffer (pH 9.0)

• Block endogenous peroxidase and non-specific binding

• Incubate with anti-CCR10 antibody at 1:20-1:200 dilution

• Apply detection system

• Counterstain, dehydrate, and mount

It is critical to include proper controls in all experimental designs to validate specific binding .

How should researchers address antibody validation for CCR10 studies?

Comprehensive validation of CCR10 antibodies should include:

• Positive and negative control tissues: Use tissues known to express CCR10 (e.g., brain tissue for mouse/rat) versus tissues with minimal expression

• Western blot validation: Confirm specificity by molecular weight (38-42 kDa expected for CCR10)

• Knockout/knockdown controls: Where possible, utilize CCR10 knockout or knockdown models to verify antibody specificity

• Cross-reactivity assessment: Test across multiple species if cross-species studies are planned

• Dilution optimization: Titrate antibodies in each experimental system to determine optimal concentration

• Blocking peptide competition: Use CCR10 peptides to compete for antibody binding as a specificity control

Studies have successfully employed CCR10 knockout models to validate antibody specificity and to investigate CCR10 function in vivo, particularly in understanding lymphocyte homing mechanisms .

How can CCR10 antibodies contribute to understanding mucosal immunity?

CCR10 antibodies have been instrumental in elucidating the role of this receptor in mucosal immune responses. Landmark studies utilizing CCR10-deficient mouse models have demonstrated that:

• CCR10 is critical for efficient localization and accumulation of IgA antibody-secreting cells (ASCs) to the lactating mammary gland

• CCR10-mediated recruitment varies dramatically between different mucosal tissues

• IgA ASC accumulation in the gastrointestinal tract is minimally impacted in CCR10-deficient mice, suggesting tissue-specific mechanisms

Researchers can employ CCR10 antibodies in immunohistochemistry to map receptor expression across mucosal tissues, in flow cytometry to quantify receptor-expressing cell populations, and in functional blocking studies to assess the impact of receptor inhibition on cellular trafficking in different experimental contexts.

What methodological approaches can be used to study CCR10 knockout models?

The development of CCR10 knockout models has been critical for understanding CCR10 function in vivo. Key methodological considerations when working with these models include:

• Generation strategy: CCR10 knockout models have been generated by replacing the first ATG codon of CCR10 exon I with an EGFP/NeoR cassette using homologous recombination in embryonic stem cells

• Genotype confirmation: Southern blot analysis of genomic DNA digested with appropriate restriction enzymes (e.g., SacI) and hybridized with labeled external probes

• Knockout validation: RT-PCR using primers designed to amplify CCR10 transcript regions:

  • Sense primer: 5′-CGGAGAAACCCTTGTAGCCAG-3′

  • Anti-sense primer: 5′-GGCCAAGACTAGGCCATTGCC-3′

• Functional assessment: Analysis of IgA ASC accumulation in mucosal tissues via flow cytometry and immunohistochemistry

When designing experiments with CCR10 knockout models, researchers should consider potential compensatory mechanisms, such as altered expression of other chemokine receptors.

What strategies can address challenges in detecting low CCR10 expression?

Detecting low levels of CCR10 expression presents technical challenges that can be addressed through several methodological approaches:

• Signal amplification techniques: Consider tyramide signal amplification for IHC applications

• Enhanced detection systems: Utilize highly sensitive detection methods like chemiluminescent substrates for Western blots

• Concentration techniques: For protein samples, consider immunoprecipitation to concentrate CCR10 before detection

• Optimized fixation protocols: Test multiple fixation methods to preserve CCR10 antigenicity

• Tissue-specific protocol adjustments: As CCR10 detection has been validated in specific tissues (brain, spleen), adapt protocols when working with other tissue types

• RNA-level detection: Complement protein detection with RT-PCR or RNA-seq to confirm expression at the transcript level

Additionally, researchers should consider the dynamic regulation of CCR10 expression, which may fluctuate based on inflammatory stimuli or cellular activation states.

How does CCR10 research relate to chemokine-based diagnostics in disease settings?

While CCR10 itself has specific roles in mucosal and cutaneous immunity, research on chemokine receptors and their ligands has broader implications for disease diagnostics. Related chemokine research has revealed:

• Urinary chemokines CXCL9 and CXCL10 (which bind to CXCR3 rather than CCR10) have demonstrated diagnostic value in antibody-mediated rejection (ABMR) after transplantation

• Combined evaluation of urinary CXCL9 with donor-specific antibody analysis improved diagnostic accuracy by 73% compared to antibody analysis alone

• Chemokine detection in non-invasive samples (urine, blood) provides valuable biomarkers that complement traditional diagnostic approaches

The methodological approaches developed for studying CCR10 and its ligands can inform similar investigations into other chemokine receptor systems with diagnostic potential.

What are the key considerations when designing studies to investigate CCR10 in pathological contexts?

When investigating CCR10 in disease settings, researchers should consider:

• Control cohort selection: Include appropriate control populations to account for baseline variations in CCR10 expression

• Tissue-specific expression patterns: Recognize that CCR10 function varies dramatically between tissue sites, necessitating site-specific investigation

• Integration with other biomarkers: As demonstrated with chemokines in transplant rejection, combined analysis with other biomarkers may provide enhanced diagnostic value

• Temporal considerations: Account for dynamic changes in CCR10 expression over disease progression

• Mechanistic validation: Complement observational studies with functional investigations using knockout models or blocking antibodies

• Multi-omics approach: Integrate antibody-based protein detection with transcriptomic and proteomic analyses for comprehensive understanding

For clinical translation, researchers should validate findings across multiple cohorts and consider the standardization of detection protocols to ensure reproducibility.

How can CCR10 antibodies be utilized in immunotherapy research?

Emerging research directions for CCR10 antibodies in immunotherapy include:

• Target validation: Using CCR10 antibodies to validate the receptor as a therapeutic target in specific disease contexts

• Immune cell trafficking modulation: Investigating how blocking CCR10 might redirect immune cells away from specific tissue sites

• Development of therapeutic antibodies: Utilizing research-grade antibodies as starting points for developing therapeutic candidates

• Biomarker identification: Exploring CCR10 expression as a potential biomarker for patient stratification in immunotherapy trials

• Combination therapy assessment: Investigating how CCR10 blockade might complement other immunotherapeutic approaches

Research on other chemokine receptors has demonstrated the potential value of this approach, as suggested by the diagnostic utility of chemokines in transplantation settings .

What novel methodological approaches are emerging for studying CCR10 biology?

Innovative approaches for investigating CCR10 biology include:

• CRISPR-based genome editing: More precise generation of receptor mutations beyond traditional knockout models

• Single-cell analysis: Examination of CCR10 expression heterogeneity within seemingly homogeneous cell populations

• In vivo imaging: Development of labeled antibodies or ligands for tracking CCR10-expressing cells in living organisms

• Receptor-ligand interaction studies: Advanced biophysical methods to characterize binding kinetics and structural determinants

• Humanized mouse models: Generation of models expressing human CCR10 for more translatable research

• Systems biology approaches: Integration of CCR10 signaling within broader chemokine network models

These methodological innovations promise to provide deeper insights into CCR10 biology and potential therapeutic applications.