C2orf15 Antibody

What is C2orf15 and what is currently known about its molecular characteristics?

C2orf15 is identified as an uncharacterized protein encoded by the chromosome 2 open reading frame 15 gene in humans. It is also referenced by the protein designation CB015_HUMAN with NCBI Gene ID 150590 . Despite being identified as a protein-coding sequence, C2orf15's specific cellular functions remain largely uncharacterized. Current bioinformatic analyses suggest extensive functional associations, with data indicating 2,447 potential biological interactions spanning 6 different categories . Research efforts continue to elucidate its structure, localization, and physiological relevance.

How is C2orf15 expression distributed across different tissue types?

Expression analysis through resources like the Allen Brain Atlas indicates tissue-specific expression patterns of C2orf15, particularly in brain tissues . This suggests potential neurological functions that warrant further investigation. Researchers should consider designing tissue-specific experiments when studying C2orf15, as expression levels may vary significantly between different anatomical regions and cell types. Additional expression data from other tissues beyond the nervous system remains limited, highlighting an area requiring further research.

What criteria should guide C2orf15 antibody selection for specific experimental applications?

When selecting C2orf15 antibodies, researchers should evaluate multiple parameters:

For uncharacterized proteins like C2orf15, validation documentation becomes particularly critical to ensure experimental reliability.

What rigorous validation approaches ensure antibody specificity for C2orf15?

Given C2orf15's uncharacterized nature, rigorous validation is essential:

• Molecular validation: Western blot analysis comparing wild-type samples with C2orf15 knockout/knockdown models to confirm band specificity at the expected molecular weight

• Cross-reactivity assessment: Testing against recombinant C2orf15 protein (such as ABIN2714092 from HEK-293 cells) alongside other proteins with sequence homology

• Epitope mapping: Determining precise binding regions to predict potential cross-reactivity with structurally similar proteins

• Cell-type specificity: Validating consistent staining patterns across multiple cell types known to express C2orf15 based on transcriptomic data

• Peptide competition assays: Pre-incubating antibody with immunizing peptide to demonstrate signal specificity

These validation approaches are particularly important for uncharacterized proteins where reference standards may be limited.

How should Western blotting protocols be optimized specifically for C2orf15 detection?

Western blotting for C2orf15 requires careful optimization:

• Sample preparation: Use RIPA buffer with protease inhibitor cocktail; sonicate briefly to ensure complete protein extraction from relevant cellular compartments

• Gel selection: Begin with 10-12% polyacrylamide gels based on the predicted molecular weight of C2orf15

• Transfer parameters: For potentially hydrophobic proteins like C2orf15, consider:

  • Extended transfer times (90-120 minutes)

  • Addition of 0.1% SDS to transfer buffer to facilitate protein movement

  • PVDF membranes for better protein retention

• Antibody conditions: For antibodies like ABIN7174820 :

  • Initial dilution range: 1:500-1:1000

  • Extended incubation (overnight at 4°C)

  • TBS-T with 5% BSA as diluent

  • Include positive control samples based on expression database information

• Detection optimization: For potentially low-abundance proteins:

  • Use high-sensitivity ECL substrates

  • Consider signal amplification systems

  • Optimize exposure times systematically

Researchers should validate these parameters empirically with each specific C2orf15 antibody.

What methodological considerations apply when using C2orf15 antibodies for immunohistochemistry applications?

For IHC applications with C2orf15 antibodies (ABIN7174820 or LS-C378304) :

• Tissue preparation:

  • For formalin-fixed paraffin-embedded samples, test multiple antigen retrieval methods (citrate pH 6.0, EDTA pH 9.0, and enzymatic approaches)

  • For frozen sections, optimize fixation conditions (4% PFA timing)

• Blocking strategy:

  • Include dual blocking steps: peroxidase block (3% H₂O₂) followed by protein block

  • For biotin-conjugated antibodies like LS-C378304 , include avidin/biotin blocking steps

• Antibody optimization:

  • Test concentration gradient (1:50 to 1:500)

  • Compare incubation conditions (1h room temperature vs. overnight 4°C)

  • For biotin-conjugated antibodies, optimize streptavidin-HRP dilution

• Signal development:

  • Time DAB development precisely to avoid background

  • Consider tyramide signal amplification for low-abundance targets

• Counterstaining:

  • Adjust hematoxylin timing to maintain visibility of potentially weak C2orf15 signals

What technical strategies enhance sensitivity and specificity in ELISA assays for C2orf15?

For developing ELISA assays using C2orf15 antibodies (ABIN7174820, ABIN7174823, LS-C378304) :

• Assay format determination:

  • For sandwich ELISA: Use two antibodies recognizing different epitopes

  • For competitive ELISA: Use recombinant C2orf15 protein (ABIN2714092) as competitor

• Protocol optimization:

  • Coating concentration: Test range from 1-10 μg/ml

  • Blocking buffer: Compare BSA-, casein-, and commercial blocking reagents

  • Sample diluent: Evaluate additives to minimize matrix effects

• Reagent titration:

  • Primary antibody: Test dilution series (typically 1:100 to 1:5000)

  • For biotin-conjugated antibodies like LS-C378304 , optimize streptavidin-HRP concentration

  • Substrate reaction timing: Develop standardized endpoint determination

• Validation parameters:

  • Establish limit of detection using recombinant protein

  • Determine assay precision (intra/inter-assay CV < 15%)

  • Spike-and-recovery experiments to assess matrix effects

How can C2orf15 antibodies contribute to protein-protein interaction studies?

C2orf15 antibodies enable multiple approaches to investigate protein-protein interactions:

• Co-immunoprecipitation strategies:

  • Forward approach: Immunoprecipitate with C2orf15 antibody and identify binding partners by mass spectrometry

  • Reverse approach: Immunoprecipitate with antibodies against predicted interactors and probe for C2orf15

  • Crosslinking optimization to capture transient interactions

• Proximity labeling coupled with immunoprecipitation:

  • BioID or APEX2 fusion proteins validated with C2orf15 antibodies

  • Use C2orf15 antibodies to confirm proximity labeling results by orthogonal methods

• Microscopy-based interaction analysis:

  • Proximity ligation assay (PLA) using C2orf15 antibodies paired with antibodies against putative interactors

  • FRET-based approaches with appropriate controls

• Validation of interactions:

  • Use recombinant C2orf15 protein (ABIN2714092) for in vitro binding assays

  • Domain mapping through truncated protein constructs

These approaches can help establish C2orf15's functional network and suggest potential biological roles.

What methodological approaches enable investigation of C2orf15 in tumor biology research?

Based on antibody applications in tumor research , C2orf15 investigation may include:

• Expression profiling across tumor types:

  • Tissue microarray analysis using validated C2orf15 antibodies

  • Correlation with clinical parameters and patient outcomes

  • Comparison with matched normal tissues

• Functional characterization:

  • siRNA/shRNA knockdown validated with C2orf15 antibodies

  • Phenotypic assays (proliferation, migration, invasion)

  • Signaling pathway analysis through phosphoprotein detection

• Tumor microenvironment interactions:

  • Multiplex immunofluorescence to examine C2orf15 expression in relation to immune cell markers

  • Analysis of B cell infiltration patterns in relation to C2orf15 expression

  • Co-culture experiments with validation by C2orf15 antibody staining

• Potential therapeutic applications:

  • Evaluate C2orf15 as target for antibody-based therapeutics if tumor-specific expression is confirmed

  • Consider antibody-superantigen fusion approaches similar to those described for other targets

How should experimental design address contradictory findings in C2orf15 expression or function?

When encountering contradictory results regarding C2orf15:

• Antibody validation reconciliation:

  • Test multiple antibodies against different epitopes

  • Confirm specificity using genetic approaches (CRISPR knockout)

  • Correlate protein detection with mRNA expression data

• Context-dependent analysis:

  • Systematically evaluate expression across different:

    • Cell types and differentiation states

    • Tissue origins and microenvironments

    • Experimental conditions (stress, nutrient availability, etc.)

• Isoform-specific investigation:

  • Design experiments to distinguish potential splice variants

  • Use antibodies with mapped epitopes to differentiate isoforms

  • Correlate with RNA-seq data to identify expression patterns

• Technical parameter assessment:

  • Standardize extraction methods to ensure consistent protein recovery

  • Compare fixation protocols that may affect epitope accessibility

  • Document lot-to-lot antibody variation

What specific troubleshooting approaches apply to weak or absent C2orf15 signal detection?

When encountering detection challenges with C2orf15 antibodies:

• Sample-related factors:

  • Verify C2orf15 expression levels in chosen samples using transcript data

  • Optimize protein extraction based on predicted cellular localization

  • Consider enrichment approaches (subcellular fractionation, immunoprecipitation)

• Antibody-specific optimization:

  • Test concentration range beyond standard recommendations

  • Evaluate multiple antibodies (ABIN7174820, LS-C378304)

  • For biotin-conjugated antibodies, confirm detection system functionality

• Protocol adjustments:

  • Western blot: Extend exposure times; consider enhanced chemiluminescence reagents

  • IHC: Optimize antigen retrieval; implement signal amplification systems

  • ELISA: Increase sample concentration; adjust incubation times

• Positive control inclusion:

  • Use recombinant C2orf15 protein (ABIN2714092) as technical control

  • Include samples with documented C2orf15 expression based on database evidence

• Systematic documentation:

  • Maintain comprehensive records of optimization attempts

  • Document reagent lots and experimental conditions

  • Consider temperature, incubation time, and buffer composition variations

How can non-specific binding be distinguished from genuine C2orf15 signal?

To differentiate specific from non-specific signals:

• Validation controls:

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

  • Genetic validation: Compare signal in wild-type vs. C2orf15 knockdown/knockout samples

  • Isotype controls: Use matched concentration of non-targeting antibody

• Signal pattern analysis:

  • Compare observed cellular/tissue distribution with RNA expression databases

  • Evaluate consistency of signal localization across different detection methods

  • Assess molecular weight precision in Western blots

• Cross-antibody validation:

  • Compare signal patterns using antibodies targeting different C2orf15 epitopes

  • Correlate signals across different applications (WB, IHC, ELISA)

• Technical refinements:

  • Optimize blocking conditions specifically for each application

  • Increase stringency of wash steps incrementally

  • For biotin-conjugated antibodies like LS-C378304 , implement additional blocking steps

• Quantitative assessment:

  • Establish signal-to-noise ratios for objective comparison

  • Document pattern consistency across technical and biological replicates

How might C2orf15 antibodies contribute to understanding tissue-specific expression patterns?

C2orf15 antibodies can advance tissue expression mapping through:

• Comprehensive tissue profiling:

  • Systematic IHC analysis across tissue types using validated antibodies

  • Correlation with data from Allen Brain Atlas and other expression databases

  • Single-cell resolution analysis in heterogeneous tissues

• Developmental expression mapping:

  • Analysis across embryonic, postnatal, and adult tissues

  • Correlation with differentiation markers

  • Temporal regulation assessment

• Comparative analysis methods:

  • Multi-omics correlation: Integrate antibody-based protein detection with transcriptomic and proteomic datasets

  • Cross-species comparison using antibodies with appropriate reactivity

  • Standardized quantification approaches for objective comparison

• Pathological state evaluation:

  • Expression analysis in disease models

  • Correlation with pathological markers

  • Potential biomarker assessment

What methodological approaches can characterize post-translational modifications of C2orf15?

To investigate PTMs of C2orf15:

• Modification-specific detection strategies:

  • Phosphorylation: Use phosphatase treatments as controls; combine with phospho-specific antibodies if available

  • Glycosylation: Employ enzymatic deglycosylation followed by mobility shift analysis

  • Ubiquitination: Use proteasome inhibitors to enhance detection

• Mass spectrometry validation:

  • Immunoprecipitate C2orf15 using available antibodies (ABIN7174820, LS-C378304)

  • Analyze by LC-MS/MS to identify modification sites

  • Compare modifications across different cellular contexts

• Functional correlation:

  • Assess modification changes in response to stimuli

  • Correlate with protein localization changes

  • Develop modification-specific antibodies if significant sites are identified

• Technical considerations:

  • Optimize sample preparation to preserve labile modifications

  • Include appropriate inhibitors during extraction

  • Validate findings with multiple methodological approaches

How can C2orf15 antibodies be integrated with emerging single-cell analysis technologies?

C2orf15 antibodies can enhance single-cell analyses through:

• Mass cytometry (CyTOF) integration:

  • Metal-conjugate C2orf15 antibodies for multiplexed analysis

  • Combine with lineage markers for cell type-specific expression

  • Correlate with functional markers in heterogeneous populations

• Spatial transcriptomics validation:

  • Combine antibody-based protein detection with spatial RNA analysis

  • Validate transcriptomic findings at protein level

  • Assess subcellular localization in tissue context

• Microfluidic applications:

  • Antibody-based sorting of C2orf15-expressing cells

  • On-chip immunoassays for limited samples

  • Integration with single-cell proteomics

• Technical optimization requirements:

  • Minimize background in low-input samples

  • Validate antibody performance in multiplexed formats

  • Develop standardized quantification approaches

These integrated approaches can provide comprehensive understanding of C2orf15 biology at unprecedented resolution.