CORO1C Antibody

What are the validated applications for CORO1C antibodies?

CORO1C antibodies have been validated for multiple experimental applications with specific optimal dilution ranges for each method:

It is recommended to optimize these dilutions for your specific experimental system . For immunohistochemistry applications with CORO1C antibodies, antigen retrieval with TE buffer (pH 9.0) is suggested, though citrate buffer (pH 6.0) can serve as an alternative .

How do I validate the specificity of a CORO1C antibody?

Antibody specificity validation should include multiple approaches:

• Molecular weight confirmation: CORO1C has a calculated molecular weight of 53 kDa but typically appears at approximately 57 kDa on SDS-PAGE gels .

• Cross-reactivity testing: Use a panel of cell lines or tissues with known CORO1C expression levels. The antibody should specifically recognize CORO1C but not other coronin family members like Coro1A or Coro1B .

• Knockdown/knockout validation: Generate CORO1C knockdown/knockout cells and confirm the loss of signal. Research groups have developed monoclonal antibodies against CORO1C that do not cross-react with other coronins, which can be used as positive controls .

• Subcellular localization confirmation: In properly functioning cells, CORO1C should strongly localize to lamellipodia and vesicular structures, showing similar localization patterns to the Arp2/3 complex .

What controls should be included when using CORO1C antibodies?

For rigorous experimental design with CORO1C antibodies, include:

• Positive controls: Cell lines with confirmed CORO1C expression, such as SGC-7901, HeLa, or SH-SY5Y cells .

• Negative controls:

  • Primary antibody omission

  • Isotype control (Rabbit IgG for polyclonal antibodies)

  • CORO1C-depleted samples (via siRNA knockdown or CRISPR knockout)

• Specificity controls: When examining lamellipodial localization, treatment with Arp2/3 inhibitors (e.g., CK666) should cause CORO1C to become diffuse in the cytoplasm, confirming proper antibody behavior .

• Loading/staining controls: Standard housekeeping proteins for western blots; nuclear counterstains for immunohistochemistry.

How can CORO1C antibodies be used to study actin dynamics and cellular migration?

CORO1C antibodies enable sophisticated investigations of actin cytoskeleton regulation:

• Lamellipodial dynamics: CORO1C strongly co-localizes with Arp2/3-branched actin networks. Using CORO1C antibodies in conjunction with live-cell imaging can reveal how this protein regulates lamellipodial protrusion dynamics .

• Co-operative binding analysis: CORO1C exhibits unique co-operative binding to F-actin with a Hill coefficient of approximately 3.922, unlike other coronin family members (Coro1A and Coro1B) that bind non-cooperatively with Hill coefficients of ~1.0 . Antibodies can help characterize these binding properties through co-sedimentation assays.

• Migration assays: CORO1C depletion leads to altered lamellipodial protrusion dynamics due to increased branched actin density and reduced actin turnover, resulting in defective cell migration . Antibodies can confirm knockdown efficiency and examine compensatory mechanisms.

• Contour-based quantification: Advanced image analysis approaches using CORO1C immunofluorescence can determine the average fluorescence intensity along contour edges, providing quantitative measurements of leading-edge localization .

What are the methodological considerations when studying CORO1C mutants?

When investigating CORO1C mutations and their effects:

How can CORO1C antibodies be utilized to investigate cancer progression mechanisms?

CORO1C antibodies are valuable tools for cancer research:

• Tissue microarray (TMA) analysis: CORO1C expression can be quantified in cancer tissues using the semi-quantitative H-score method:

  • Score 0: negative expression

  • Score 1: weakly positive staining

  • Score 2: moderately positive staining

  • Score 3: strongly positive staining

  Final staining scores are calculated by multiplying each intensity score by the percentage of cells at that intensity and summing the results (maximum possible score: 300) .

• Correlation with clinical parameters: CORO1C expression has been significantly associated with:

  • Histological type

  • Lymph node metastasis

  • Distant metastasis

  • AJCC stage

  • Venous invasion

  • Perineural invasion

• Signaling pathway analysis: CORO1C antibodies can help investigate the PI3K/AKT signaling pathway activation, which mediates the tumor-promoting role of CORO1C .

• EMT marker correlation: Combine CORO1C antibody staining with epithelial-mesenchymal transition markers to understand mechanistic relationships in cancer progression .

What are common issues when using CORO1C antibodies and how can they be addressed?

When working with CORO1C antibodies, researchers may encounter:

• High background in immunostaining:

  • Solution: Optimize blocking conditions (try 5% BSA or 5% normal serum from the secondary antibody host species)

  • Increase washing steps duration and number

  • Further dilute the primary antibody (try 1:800 for IF applications)

• Weak or no signal in Western blots:

  • Solution: Ensure proper sample preparation (CORO1C is observed at 57 kDa)

  • Optimize protein loading (30-50 μg total protein recommended)

  • Decrease antibody dilution (try 1:500)

  • Consider enhanced chemiluminescence detection systems

• Non-specific bands:

  • Solution: Validate specificity using CORO1C knockout/knockdown samples

  • Use more stringent washing conditions

  • Consider monoclonal antibodies with confirmed specificity

• Variable staining intensity across samples:

  • Solution: Standardize fixation protocols

  • For IHC, ensure consistent antigen retrieval (recommended: TE buffer pH 9.0)

  • Include positive control samples in each experiment

How should CORO1C antibodies be stored and handled for optimal performance?

To maintain antibody performance:

• Storage conditions: Store at -20°C in the buffer provided (typically PBS with 0.02% sodium azide and 50% glycerol, pH 7.3) .

• Stability: Most CORO1C antibodies remain stable for one year after shipment when stored properly .

• Aliquoting: For antibodies stored at -20°C, aliquoting is typically unnecessary, though it can prevent freeze-thaw cycles for frequently used antibodies .

• Working solution preparation: Dilute only the amount needed for immediate use in appropriate buffer containing 1-5% BSA or normal serum.

• Shipping and temporary storage: Can withstand ambient temperatures during shipping, but should be stored at recommended temperatures immediately upon receipt.

How does CORO1C function compare to other coronin family members?

Key differences between CORO1C and other coronins include:

• Binding properties:

  • CORO1C exhibits positive co-operative binding to F-actin (Hill coefficient ~3.922)

  • CORO1A and CORO1B bind non-cooperatively (Hill coefficient ~1)

• Structural features:

  • CORO1C contains a unique second actin-binding site that enhances binding activity

  • Mutations in either binding site (R28D or 2×KE) block co-operative binding

• Functional redundancy:

  • CORO1B and CORO1C show some overlapping functions in regulating lamellipodia dynamics

  • For accurate functional studies, simultaneous depletion of both proteins is recommended to avoid compensation effects

• Localization patterns:

  • All coronin family members localize to actin-rich structures

  • CORO1C specifically shows strong lamellipodial and vesicular localization similar to the Arp2/3 complex

  • CORO1C localization is dependent on Arp2/3 activity, as inhibition with CK666 causes diffuse cytoplasmic distribution

What are the emerging research directions for CORO1C antibodies in cancer research?

Current and future research applications include:

• Prognostic biomarker development: CORO1C expression correlates with poor prognosis in colorectal cancer patients, suggesting potential use as a prognostic marker .

• Therapeutic target identification: Given CORO1C's role in promoting metastasis via the PI3K/AKT pathway, combined targeting approaches might be investigated .

• Multi-omics integration: Correlating CORO1C protein expression (detected by antibodies) with transcriptomic and proteomic datasets to identify regulatory networks.

• Microenvironment interactions: Investigating how CORO1C-expressing cancer cells interact with stromal components using co-culture systems and multiplexed immunostaining.

• Drug response prediction: Testing whether CORO1C expression levels (detected via antibodies) correlate with response to cytoskeleton-targeting therapies or PI3K/AKT inhibitors .

How can CORO1C antibodies be used in combination with other molecular techniques?

Integrating CORO1C antibodies with complementary methodologies enhances research depth:

• Live-cell imaging combined with fixed-cell antibody staining: Track dynamics in live cells (using fluorescent-tagged constructs), then fix and immunostain the same cells to correlate with endogenous protein localization .

• Proximity ligation assays (PLA): Identify protein-protein interactions between CORO1C and binding partners like Trop2, which has been shown to interact with CORO1C in colorectal cancer .

• Super-resolution microscopy: Combine with CORO1C antibodies to visualize nanoscale organization within lamellipodia and other actin-rich structures.

• CRISPR/Cas9 gene editing: Create precise mutations in endogenous CORO1C and use antibodies to verify protein expression or assess localization changes .

• Tissue microarrays with multiplex immunostaining: Simultaneously detect CORO1C and other markers (e.g., EMT markers, PI3K/AKT pathway components) to build comprehensive protein expression profiles in cancer progression .