CCDC184 Antibody

What is CCDC184 protein and what are its known biological functions?

CCDC184 (Coiled-coil domain-containing protein 184) is a human protein with UniProt ID Q52MB2, also known by alternative names including C12orf68 (Chromosome 12 open reading frame 68) and LOC387856 . The protein contains coiled-coil structural motifs that typically facilitate protein-protein interactions. While comprehensive functional studies on CCDC184 are still emerging in the literature, the protein's coiled-coil structure suggests potential roles in cellular structural organization, signaling pathways, or regulatory functions. The gene is located on chromosome 12 in humans, and current research indicates expression in multiple tissue types, including findings from immunohistochemistry studies showing detection in lung cancer and melanoma tissues .

What are the primary research applications for CCDC184 antibodies?

Based on validated testing parameters, CCDC184 antibodies have demonstrated utility in several key research applications:

Notably, CCDC184 antibodies have been successfully applied in immunohistochemical analysis of human melanoma and lung cancer tissues, suggesting particular relevance for cancer research applications . The diversity of validated applications indicates the versatility of these antibodies for multiple experimental approaches in studying CCDC184 expression and function.

How should researchers select between polyclonal and monoclonal CCDC184 antibodies?

The selection between polyclonal and monoclonal CCDC184 antibodies should be guided by specific experimental requirements. Currently available CCDC184 antibodies are predominantly rabbit polyclonal antibodies . Polyclonal antibodies offer advantages for detecting CCDC184 in complex samples due to their recognition of multiple epitopes, potentially enhancing sensitivity for low-abundance targets. This multi-epitope recognition is particularly valuable when studying proteins like CCDC184 where conformational changes might occur under different experimental conditions.

For experiments requiring absolute epitope specificity or batch-to-batch consistency over extended research periods, researchers might consider exploring whether monoclonal options become available. The decision should be influenced by factors including the intended application, required sensitivity, available sample quantities, and whether the research focuses on specific domains of the CCDC184 protein.

What are the optimal storage and handling conditions for maintaining CCDC184 antibody activity?

To maintain CCDC184 antibody functionality, researchers should adhere to specific storage protocols depending on intended usage timeframes:

• Short-term storage (up to 2 weeks): Maintain at refrigerated temperatures (2-8°C) .

• Long-term storage: Store at -20°C in small aliquots to prevent repeated freeze-thaw cycles that can compromise antibody activity .

• Working solution preparation: When preparing dilutions for experiments, use sterile conditions and appropriate buffer systems containing 50% glycerol and preservatives such as 0.03% Proclin 300 to maintain stability .

For functional grade antibodies, additional precautions regarding sterility and endotoxin levels may be necessary. Storage in small aliquots is particularly critical as each freeze-thaw cycle can reduce antibody activity by approximately 10-15%. Documentation of freeze-thaw cycles and preparation dates is recommended for experimental reproducibility.

How should immunohistochemistry protocols be optimized for CCDC184 detection in different tissue types?

Optimization of immunohistochemistry protocols for CCDC184 detection requires systematic adjustment of multiple parameters:

• Antigen retrieval method: For paraffin-embedded tissues, heat-induced epitope retrieval using citrate buffer (pH 6.0) or EDTA buffer (pH 9.0) should be compared to determine optimal epitope exposure.

• Antibody dilution: Start with the manufacturer's recommended range (1:20-1:200 for IHC-P) and perform a dilution series to identify the optimal signal-to-noise ratio for your specific tissue type.

• Incubation conditions: Test both overnight incubation at 4°C and shorter incubations (1-2 hours) at room temperature to determine which provides optimal staining.

• Detection system selection: For tissues with potentially low CCDC184 expression, a high-sensitivity detection system may be required. Compare DAB-based chromogenic detection with fluorescence-based methods for specific research questions.

• Positive controls: Include melanoma or lung cancer tissue sections as positive controls, as these have demonstrated CCDC184 expression in previous studies .

The optimization should be conducted systematically, changing only one parameter at a time while documenting outcomes to establish a reproducible protocol for your specific experimental system.

What are the recommended approaches for validating CCDC184 antibody specificity?

Comprehensive validation of CCDC184 antibody specificity should incorporate multiple complementary approaches:

• Peptide competition assays: Pre-incubate the antibody with the immunogen peptide (recombinant human CCDC184 protein fragments 1-194AA) before application to demonstrate signal reduction in the presence of the specific target.

• Knockout/knockdown controls: Utilize CCDC184 knockdown or knockout cell lines as negative controls to confirm signal specificity.

• Multiple antibody validation: Compare staining patterns using antibodies targeting different epitopes of CCDC184, ideally from different manufacturers or production lots.

• Protein array testing: Cross-reference with protein array data testing the antibody against multiple human recombinant proteins to assess potential cross-reactivity .

• Western blot analysis: Confirm that the detected protein band corresponds to the expected molecular weight of CCDC184.

• Mass spectrometry validation: For definitive validation, immunoprecipitation followed by mass spectrometry can confirm the identity of the pulled-down protein.

Documentation of these validation steps is essential for publication and experimental reproducibility, particularly given the potential for inconsistencies in antibody-based experiments highlighted in pharmacogenomic research .

How do various conjugated forms of CCDC184 antibodies impact experimental design and outcomes?

CCDC184 antibodies are available in multiple conjugated formats, each optimized for specific experimental applications:

When selecting between these formats, researchers should consider:

• The sensitivity requirements of their experimental system

• The potential for background from secondary detection reagents

• Multiplexing needs (when combining with other antibodies)

• Signal amplification requirements for low-abundance targets

The conjugation chemistry can occasionally impact epitope recognition, so preliminary testing of different formats may be necessary when first establishing a new experimental system. Additionally, some applications may benefit from specific conjugates - for example, biotin conjugation may provide superior signal amplification for tissues with low CCDC184 expression.

What cross-reactivity considerations should be addressed when working with CCDC184 antibodies across species?

Understanding the cross-reactivity profile of CCDC184 antibodies is crucial for experimental design across different model systems:

Current commercial CCDC184 antibodies demonstrate variable species reactivity profiles. While some are specifically validated for human samples , others show broader reactivity including mouse models . Additionally, some CCDC184 antibody products indicate potential cross-reactivity with bovine, canine, equine, guinea pig, and rat proteins .

When planning cross-species experiments, researchers should:

• Conduct sequence homology analysis between human CCDC184 and the orthologous protein in the target species, particularly focusing on the immunogen region (for available products, this includes the 1-194AA region) .

• Perform preliminary validation experiments in the non-human species tissues before conducting full studies.

• Consider using antibodies raised against immunogens from the specific species of interest when available.

• Include appropriate positive and negative controls from each species in all experiments.

The degree of sequence conservation in the immunogen region will significantly impact cross-reactivity potential. Epitope mapping can provide valuable insights into the specific binding regions and potential cross-reactivity mechanisms.

How can researchers address inconsistent results when working with CCDC184 antibodies?

Inconsistencies in antibody-based experiments represent a significant challenge highlighted in pharmacogenomic research . For CCDC184 antibody experiments specifically, a systematic troubleshooting approach should include:

• Antibody validation reassessment: Confirm antibody specificity using methods outlined in section 2.3, particularly when using a new lot or after extended storage.

• Protocol standardization: Implement rigorous standardization of all experimental parameters including sample preparation, antibody dilution calculation methods, incubation times, and detection systems.

• Positive control inclusion: Include consistent positive controls (such as melanoma or lung cancer tissues with known CCDC184 expression) across experimental batches.

• Quantification methods: Establish consistent quantification approaches for CCDC184 signals, including appropriate thresholding and normalization strategies.

• Intra-laboratory validation: Have multiple researchers perform identical protocols to identify potential operator-dependent variables.

• Multi-technique confirmation: Validate findings using complementary techniques (e.g., confirming IHC results with Western blot or qPCR).

• Batch effect monitoring: Record and analyze potential batch effects including antibody lots, reagent preparations, and environmental variables.

When inconsistencies persist despite these measures, meta-analysis approaches combining data from multiple experiments may help identify reproducible patterns amidst experimental noise. The field of pharmacogenomics has developed sophisticated statistical approaches for addressing inconsistencies that can be adapted to antibody-based research .

What quantification approaches are most appropriate for CCDC184 expression analysis?

Quantitative analysis of CCDC184 expression requires selection of appropriate methodologies based on the experimental technique:

For immunohistochemistry:

• H-score method: Calculate intensity (0-3+) multiplied by percentage of positive cells (0-100%) for scores ranging from 0-300.

• Allred scoring: Combine proportion score (0-5) and intensity score (0-3) for a total score of 0-8.

• Digital image analysis: Utilize automated systems with consistent thresholding parameters for objective quantification.

For Western blot analysis:

• Densitometry normalized to loading controls (β-actin, GAPDH)

• Calculation of relative expression compared to standardized positive controls

For immunofluorescence:

• Mean fluorescence intensity measurements with background subtraction

• Quantification of subcellular distribution patterns

The selection of appropriate quantification methods should be guided by:

• The specific research question (presence/absence vs. expression level vs. localization)

• The sensitivity requirements of the experiment

• The availability of appropriate normalization controls

• The statistical approaches planned for downstream analysis

Documentation of the specific quantification methodology is essential for experimental reproducibility and cross-study comparisons.

How should researchers interpret CCDC184 localization patterns in subcellular compartments?

Interpretation of CCDC184 subcellular localization requires careful consideration of multiple factors:

Based on current research, CCDC184 has been detected using immunofluorescence techniques with recommended dilutions of 0.25-2 μg/mL . When analyzing subcellular localization patterns, researchers should:

• Employ co-localization studies with established subcellular markers (e.g., DAPI for nucleus, phalloidin for cytoskeleton, organelle-specific markers).

• Compare observed patterns to predicted localization based on protein sequence analysis for known localization signals (nuclear localization signals, membrane targeting sequences, etc.).

• Consider fixation and permeabilization effects on apparent localization - compare multiple fixation methods to confirm consistent patterns.

• Utilize super-resolution microscopy techniques for detailed localization studies when conventional microscopy provides ambiguous results.

• Complement imaging studies with subcellular fractionation and Western blot analysis of different cellular compartments.

Changes in CCDC184 localization under different experimental conditions or disease states may provide valuable insights into protein function. Quantitative co-localization analysis using Pearson's correlation coefficient or Manders' overlap coefficient can provide objective measures of spatial relationships with other cellular components.

What are promising research directions for elucidating CCDC184 function in normal and disease states?

Based on current knowledge and experimental capabilities with CCDC184 antibodies, several research directions show particular promise:

• Comprehensive tissue expression profiling: Leveraging validated CCDC184 antibodies for immunohistochemistry to map expression across normal human tissues and disease states, particularly expanding beyond the current studies in melanoma and lung cancer .

• Protein interaction network mapping: Using CCDC184 antibodies for co-immunoprecipitation followed by mass spectrometry to identify binding partners that could illuminate functional pathways.

• Post-translational modification characterization: Developing modification-specific antibodies to understand how phosphorylation, ubiquitination, or other modifications regulate CCDC184 function.

• Structure-function relationship studies: Correlating antibody epitope accessibility with protein structural changes under different cellular conditions.

• CCDC184 in cancer progression: Given the preliminary observations in cancer tissues , systematic studies across cancer progression stages could reveal potential biomarker applications.

These research directions would substantially advance our understanding of this relatively understudied protein and potentially reveal new therapeutic targets or diagnostic approaches.

How might emerging antibody technologies enhance CCDC184 research in the near future?

Several emerging technologies in antibody development and application hold particular promise for advancing CCDC184 research:

• Recombinant antibody technology: Development of recombinant CCDC184 antibodies with defined clonality and epitope targeting could address current inconsistency challenges by providing renewable, precisely characterized reagents.

• Nanobodies and single-domain antibodies: Smaller antibody formats could enable improved penetration in tissue sections and potentially access epitopes unavailable to conventional antibodies.

• Multiplexed immunofluorescence platforms: Integration of CCDC184 detection into multiplexed panels would allow simultaneous analysis of multiple markers, providing contextual information about CCDC184 expression in relation to cell types and signaling pathways.

• Proximity labeling techniques: Conjugation of CCDC184 antibodies with enzymes that catalyze proximity-dependent labeling could enable detailed analysis of the protein's immediate microenvironment.

• In vivo imaging applications: Development of CCDC184 antibody fragments suitable for in vivo imaging could enable tracking of expression in animal models during disease progression or treatment response.

These technologies would substantially expand the experimental toolkit available for CCDC184 research, potentially accelerating functional discoveries and translational applications.