CCDC150 Antibody

What is CCDC150 and why is it significant in research?

CCDC150 is a coiled-coil domain-containing protein that has gained research interest due to its significant up-regulation in triple-negative breast cancer. Studies have demonstrated a 19.31-fold increase in CCDC150 expression in TNBC tumors compared to adjacent tissues . The protein is closely related to cellular component organization and cytoskeletal function, making it particularly relevant for cancer migration and invasion research . CCDC150 has been identified as a potential therapeutic target due to its role in promoting TNBC migration and invasion through the TGF-β1/SMAD3 signaling pathway .

What types of CCDC150 antibodies are available for research applications?

Research-grade CCDC150 antibodies are primarily available as polyclonal antibodies, typically derived from rabbit hosts. Commercial options include unconjugated antibodies targeted against various regions of the human CCDC150 protein . These antibodies are generally suitable for applications such as ELISA and immunohistochemistry (IHC) . Some antibodies are raised against recombinant fragments of the protein, such as amino acids 1-245 or 367-461, which allows for targeted recognition of specific domains .

What is the molecular characterization of CCDC150 protein?

CCDC150 is characterized by its coiled-coil domain structure and is identified with UniProt ID Q8NCX0 and Entrez Gene ID 284992 for the human variant . The functional analysis of CCDC150 indicates its involvement in cytoskeletal organization, which directly impacts cellular migration and invasion capabilities . Bioinformatics analyses have revealed that CCDC150 expression patterns vary across different TNBC subtypes, including BL1, BL2, IM, MSL, M, and UNS, with significant implications for patient prognosis .

What are the recommended applications for CCDC150 antibodies?

Based on the available research resources, CCDC150 antibodies have been validated for several experimental applications:

For optimal results in specific research contexts, optimization of antibody concentration is recommended based on the experimental system and target tissue/cell type .

How should CCDC150 antibody validation be performed?

Proper validation of CCDC150 antibodies should follow a multi-step approach:

• Use recombinant CCDC150 protein fragments as positive controls to confirm antibody specificity .

• Perform blocking experiments by pre-incubating the antibody with a 100x molar excess of the protein control fragment for 30 minutes at room temperature before application .

• Include negative controls such as isotype-matched irrelevant antibodies to identify non-specific binding.

• Validate across multiple applications (e.g., if using for both IHC and Western blot) as specificity can vary by technique.

• When possible, confirm results with genetic approaches such as siRNA knockdown of CCDC150 to demonstrate reduction in antibody signal .

This comprehensive validation approach ensures reliable experimental outcomes and minimizes false positives in CCDC150 research.

What is the significance of ortholog cross-reactivity for CCDC150 antibodies?

Understanding cross-reactivity with orthologs is crucial when designing experiments with animal models. Commercial CCDC150 antibodies show varying degrees of cross-reactivity:

• Antibodies against amino acids 367-461 show highest sequence identity to mouse and rat orthologs at approximately 83% .

• Antibodies against the immunogen sequence containing "ISPIQNEAIC...VMNL" demonstrate approximately 74% sequence identity to mouse and rat orthologs .

This cross-reactivity information is essential when selecting antibodies for research involving multiple species or animal models, as decreased sequence homology may result in reduced binding affinity and altered experimental outcomes .

How does CCDC150 expression correlate with TNBC progression and prognosis?

Research has established significant correlations between CCDC150 expression and TNBC progression:

• CCDC150 shows a 19.31-fold up-regulation in TNBC tumors compared to adjacent normal tissues .

• Expression levels vary by TNM staging, with significant increases in stage I and stage IV compared to stage III .

• High CCDC150 expression positively correlates with poor survival outcomes in TNBC patients, as demonstrated through survival curve analyses .

• Expression varies across TNBC molecular subtypes (BL1, BL2, IM, MSL, M, and UNS) .

These findings support the potential utility of CCDC150 as both a prognostic biomarker and therapeutic target in TNBC management strategies .

What cellular mechanisms are influenced by CCDC150 in cancer cells?

CCDC150 influences several critical cellular processes in cancer progression:

• Cell Cycle Regulation: Knockdown of CCDC150 arrests cells in the G1 phase of the cell cycle .

• Apoptosis: CCDC150 silencing significantly promotes early and late apoptosis in MDA-MB-231 cells .

• Migration and Invasion: CCDC150 knockdown reduces migration, invasion, and wound-healing rates in TNBC cell lines .

• Cytoskeletal Organization: CCDC150 silencing leads to F-actin rearrangement, increased polarization coefficient, and increased cell spreading area .

• Epithelial-Mesenchymal Transition (EMT): CCDC150 knockdown increases E-cadherin (2.04-fold) and decreases N-cadherin (by 48.13%), suggesting reversal of EMT .

These mechanistic insights provide valuable contexts for interpreting antibody-based detection of CCDC150 in cancer research applications .

How can CCDC150 antibodies be used to investigate TGF-β1/SMAD3 signaling pathways?

CCDC150 has been identified as an activator of the TGF-β1/SMAD3 signaling pathway in TNBC . To investigate this connection using CCDC150 antibodies:

• Perform co-immunoprecipitation experiments using CCDC150 antibodies to identify protein-protein interactions with TGF-β1 and SMAD3 components.

• Use CCDC150 antibodies in conjunction with phospho-specific SMAD3 antibodies to assess pathway activation status through immunoblotting or immunofluorescence.

• Conduct chromatin immunoprecipitation (ChIP) assays using CCDC150 antibodies to identify potential gene regulatory functions.

• Design dual immunofluorescence experiments to evaluate co-localization of CCDC150 with TGF-β1 signaling components.

These approaches can elucidate the molecular mechanisms by which CCDC150 influences TGF-β1/SMAD3 signaling in cancer progression .

What strategies can be employed to improve CCDC150 antibody specificity for challenging applications?

For applications requiring enhanced specificity:

• Pre-absorption techniques: Pre-incubate the CCDC150 antibody with a 100x molar excess of the protein control fragment for 30 minutes at room temperature before application .

• Epitope mapping: Characterize the specific binding regions to select antibodies that target unique epitopes with minimal cross-reactivity.

• Biophysics-informed modeling: Employ computational approaches similar to those used in antibody engineering to identify potential cross-reactive epitopes and select antibodies with optimal specificity profiles .

• Custom antibody design: Consider developing custom antibodies targeting specific epitopes unique to CCDC150 to minimize cross-reactivity with related proteins .

These strategies can significantly enhance experimental outcomes in applications where standard commercial antibodies may produce ambiguous results.

How can CCDC150 antibodies be integrated into multiplexed imaging approaches?

Advanced multiplexed imaging with CCDC150 antibodies requires careful consideration of:

• Antibody conjugation: Select fluorophores or enzyme labels with minimal spectral overlap when combining with other antibodies.

• Sequential staining protocols: Develop protocols for sequential application and elution of antibodies for highly multiplexed analyses.

• Validation of multiplexed signals: Confirm that antibody performance is not compromised in multiplexed formats through appropriate controls.

• Spatial context analysis: Analyze CCDC150 expression in the context of cytoskeletal markers and TGF-β pathway components for comprehensive understanding of spatial relationships.

These approaches enable sophisticated analyses of CCDC150 in complex tissue architectures and cellular contexts.

What are common challenges when using CCDC150 antibodies in clinical samples?

Researchers commonly encounter several challenges when applying CCDC150 antibodies to clinical specimens:

• Tissue heterogeneity: CCDC150 expression varies significantly across TNBC subtypes, requiring careful sample characterization .

• Fixation effects: Formalin fixation can mask epitopes, potentially requiring optimization of antigen retrieval methods.

• Specificity concerns: The 74-83% sequence identity with mouse and rat orthologs may introduce cross-reactivity issues in studies involving mixed human-animal components .

• Background signal: Coiled-coil domains share structural similarities with other proteins, potentially contributing to non-specific binding.

To address these challenges, researchers should incorporate appropriate positive and negative controls, and validate antibody specificity in each experimental context .

How should researchers interpret discrepancies in CCDC150 detection across different antibodies?

When facing discrepant results using different CCDC150 antibodies:

• Epitope differences: Different antibodies target distinct regions of CCDC150, which may be differentially accessible in certain experimental contexts or may be affected by post-translational modifications.

• Antibody class variations: Polyclonal antibodies may recognize multiple epitopes, while monoclonal antibodies target specific epitopes with higher specificity but potentially lower sensitivity.

• Protocol optimization: Each antibody may require specific conditions for optimal performance, including dilution, incubation time, and buffer composition.

• Confirmation approaches: Use orthogonal methods such as RNA expression analysis or alternative antibodies targeting different epitopes to resolve discrepancies.

How might CCDC150 antibodies contribute to developing targeted therapies for TNBC?

CCDC150 antibodies could facilitate therapeutic development through:

• Target validation: Confirming CCDC150's role in TNBC progression and metastasis through precise localization and interaction studies .

• Drug screening: Developing high-throughput screening assays using CCDC150 antibodies to identify compounds that modulate its expression or function.

• Biomarker development: Establishing standardized immunohistochemical protocols for patient stratification based on CCDC150 expression levels .

• Antibody-drug conjugates: Exploring the potential for therapeutic antibodies against cell-surface exposed domains of CCDC150, if present.

• Combination therapy approaches: Investigating CCDC150 in combination with gradient rotating magnetic field (RMF) exposure, which has shown promising results in suppressing TNBC tumor growth and metastasis .

These applications highlight the translational potential of CCDC150 research beyond basic science investigations .

What emerging antibody technologies might enhance CCDC150 research?

Emerging technologies that could advance CCDC150 antibody applications include:

• Nanobody development: Single-domain antibodies derived from camelid heavy-chain-only antibodies offer advantages in size, stability, and tissue penetration for certain applications .

• Phage display techniques: Custom antibody generation through phage display allows selection of antibodies with precise specificity profiles .

• Computational antibody design: Biophysics-informed modeling approaches enable the design of antibodies with customized specificity for closely related epitopes .

• BCR repertoire mining: Searching human B-cell receptor repertoires for antibodies targeting specific CCDC150 epitopes may yield highly specific reagents .

These emerging approaches could address current limitations in CCDC150 antibody research and expand the available toolkit for investigating this promising cancer target.