CKX7 Antibody

What is Cytokeratin 7 and what is its biological function?

Cytokeratin 7 (CK7, K7) is a type II intermediate filament protein encoded by the KRT7 gene located on chromosome 12q12-q13. It forms an essential part of the cytoskeleton in glandular and transitional epithelia. Beyond its structural role, CK7 has been shown to block interferon-dependent interphase and stimulate DNA synthesis in cells. It is also involved in the translational regulation of human papillomavirus type 16 E7 mRNA (HPV16 E7) . As a member of the keratin family, CK7 contributes to cellular mechanical stability and integrity while participating in various signaling pathways that regulate cell proliferation, differentiation, and death.

What are the key expression patterns of CK7 in normal and neoplastic tissues?

CK7 demonstrates specific expression patterns that are valuable for diagnostic purposes:

Normal Tissue Expression:

• Ductal epithelia (pancreatic ducts, bile ducts)

• Glandular epithelia

• Transitional epithelium

• Respiratory epithelium

• Female genital tract epithelium

Positive Neoplastic Tissues:

• Breast carcinoma

• Lung adenocarcinoma

• Ovarian carcinoma

• Pancreatic and biliary tract carcinomas

• Thyroid tumors (adenoma and carcinoma)

• Cervical adenocarcinoma and squamous cell carcinoma

• Salivary gland carcinoma (93%)

Negative Neoplastic Tissues:

• Colorectal carcinoma (typically CK7-/CK20+)

• Hepatocellular carcinoma

• Prostatic adenocarcinoma

• Clear cell renal cell carcinoma

• Most squamous cell carcinomas (with exceptions)

How does CK7 antibody aid in the differentiation between various tumor types?

CK7 antibody plays a critical role in tumor classification when used either alone or in combination with other markers. The most common diagnostic application involves CK7/CK20 immunophenotyping, which helps distinguish between primary and metastatic adenocarcinomas. For example:

• CK7+/CK20-: Typically seen in breast, lung, thyroid, endometrial, and ovarian carcinomas

• CK7-/CK20+: Characteristic of colorectal carcinoma and Merkel cell carcinoma

• CK7+/CK20+: Common in pancreaticobiliary, gastric, and some urothelial carcinomas

• CK7-/CK20-: Seen in hepatocellular carcinoma, prostate adenocarcinoma, and most squamous cell carcinomas

This profiling system is particularly valuable when investigating tumors of uncertain origin, as it can narrow down potential primary sites significantly.

How do different CK7 antibody clones compare in terms of specificity and sensitivity?

Various CK7 antibody clones exhibit different performance characteristics that can impact research results:

Mouse Monoclonal OV-TL 12/30:
This traditional clone has been widely used and serves as a reference standard in many laboratories. It provides consistent staining of CK7-positive tissues but may show lower intensity than newer clones.

Rabbit Monoclonal BC1:
This more recently developed clone has demonstrated superior performance characteristics compared to traditional mouse monoclonals. Research indicates that BC1 provides enhanced staining intensity, particularly in lung and breast cancer specimens, while maintaining equivalent specificity . The BC1 clone has also shown improved staining of cellular debris, which can be valuable in certain diagnostic contexts .

Researchers should select the appropriate clone based on their specific application, tissue type, and detection system.

What are the prognostic implications of CK7 expression in different cancer types?

CK7 expression patterns have been associated with prognostic outcomes in several tumor types:

Colorectal Cancer:

Esophageal Squamous Cell Carcinoma:

Intrahepatic Cholangiocarcinoma:

• CK7/CK19 index is an independent adverse prognostic factor for postoperative survival

Cervical Lesions:

• Expression in cervical low-grade squamous intraepithelial lesion (LSIL/CIN 1) is associated with higher rates of subsequent high-grade lesions, although the clinical utility may be limited by the low magnitude of risk difference and interpretive variability

These findings suggest that CK7 expression analysis can provide valuable prognostic information beyond its diagnostic utility, potentially influencing treatment decisions and patient management.

How can CK7 antibody be integrated into multiplex immunohistochemical assays?

Multiplex immunohistochemistry combining CK7 with other markers has emerged as a powerful approach for comprehensive tumor characterization:

CK7/CDX2 Multiplex Assay:
Researchers have developed a 4-step double stain (multiplex) assay by combining CDX-2 with CK7 in an antibody cocktail. This approach has been successfully tested on lung, breast, and colon cancers, allowing simultaneous visualization of both markers . This multiplexing strategy enhances diagnostic efficiency by providing more information from a single tissue section.

Three 7 Markers Approach:
For renal tumors, a multiplex panel combining "three 7 markers" (CK7, Claudin-7, and CD117) significantly increases specificity for chromophobe renal cell carcinoma versus histologic mimics . This demonstrates how CK7 can be effectively integrated into comprehensive diagnostic algorithms.

Technical Considerations:

• Antibody selection is critical, as some clones may perform better in multiplex systems

• Sequential versus cocktail approaches must be optimized based on specific antibody combinations

• Visualization systems must be carefully selected to allow clear distinction between markers

• Automated platforms may offer advantages in standardization for multiplex assays

What are the optimal protocols for CK7 immunohistochemistry in different sample types?

Successful CK7 immunohistochemistry requires optimization based on sample type and preparation method:

Formalin-Fixed Paraffin-Embedded (FFPE) Tissues:

• Antigen retrieval: Heat-induced epitope retrieval using citrate buffer (pH 6.0) or EDTA buffer (pH 9.0)

• Antibody dilution: For ab53123 (polyclonal), 1:50 dilution for immunohistochemistry on paraffin sections has been validated

• Incubation conditions: Typically 30-60 minutes at room temperature or overnight at 4°C

• Detection system: Polymer-based or avidin-biotin systems are commonly used

Western Blotting:

• Protein extraction: Standard lysis buffers are effective for CK7 extraction

• Sample preparation: Reducing conditions recommended

• Antibody dilution: 1:300 dilution has been validated for ab53123

• Expected band size: 51 kDa

Immunofluorescence/Immunocytochemistry:

• Fixation: 4% paraformaldehyde for 10 minutes

• Permeabilization: 0.1% Triton X-100

• Blocking: 1% BSA in PBS

• Cell types: HeLa cells have been validated as positive controls

What controls should be used to validate CK7 antibody performance?

Proper controls are essential for ensuring the reliability of CK7 immunostaining:

Positive Tissue Controls:

• Pancreatic ducts serve as excellent positive controls for CK7 expression

• Breast carcinoma tissue provides a reliable positive control for tumor diagnostics

• HepG2 and HeLa cell extracts have been validated for western blotting and immunocytochemistry, respectively

Negative Tissue Controls:

• Colorectal tissue generally serves as a reliable negative control

• Prostatic tissue typically shows negative staining

Peptide Controls:

• Immunizing peptide blocking experiments can confirm antibody specificity

• This approach has been demonstrated with ab53123, where the specific immunizing peptide successfully blocked antibody binding in western blot and immunohistochemistry applications

Preanalytical Variables:
Researchers should standardize fixation time, processing protocols, and storage conditions to ensure consistent immunoreactivity across specimens.

How should researchers interpret variable or focal CK7 expression in tumor samples?

Interpretation of variable CK7 expression requires consideration of several factors:

Threshold Definitions:

• Inter-laboratory variability exists in defining positivity thresholds (e.g., >1% vs. >10% positive cells)

• Researchers should establish and clearly report their positivity thresholds in publications

Staining Patterns:

• Distribution (focal vs. diffuse)

• Intensity (weak, moderate, strong)

• Subcellular localization (membranous, cytoplasmic)

Biological Significance:

• Focal positivity may indicate tumor heterogeneity

• In some contexts, even limited CK7 expression may have diagnostic or prognostic relevance

• Tumor grade, differentiation state, and microenvironment may influence expression patterns

Challenging Scenarios:

• Clear cell renal cell carcinoma typically presents as CK7 negative, but immunoreactivity can be variable, with low-grade clear cell RCC showing positivity rates of 60-93%

• Squamous cell carcinomas are generally CK7 negative, but certain subtypes (cervical, some head and neck, and poorly differentiated cutaneous variants) can express focal-to-partial CK7 positivity

How can CK7 antibody help distinguish primary from metastatic tumors?

CK7 antibody serves as a valuable tool in distinguishing primary from metastatic tumors, particularly in challenging cases:

Lung Nodules:

• CK7+/TTF1+ profile supports primary lung adenocarcinoma

• CK7-/CK20+/CDX2+ profile suggests metastatic colorectal carcinoma

• This distinction is critical for treatment planning, as therapeutic approaches differ significantly between primary and metastatic disease

Ovarian Masses:

• CK7+/CK20- pattern in ovarian mucinous carcinomas suggests primary ovarian origin

• CK7-/CK20+ pattern suggests metastatic colorectal carcinoma to the ovary

• CK7+/CK20+ pattern requires additional markers (e.g., CDX2, PAX8) for definitive classification

Liver Lesions:

• CK7+/CK20-/HepPar1- pattern in liver lesions suggests metastatic breast or lung carcinoma

• CK7-/CK20+ pattern suggests metastatic colorectal carcinoma

Specific Diagnostic Challenges:

• Adenocarcinoma of distal esophagus vs. proximal stomach: Variable CK7/CK20 patterns, but CK7+/CK20- pattern favors esophageal origin

• Chromophobe renal cell carcinoma (diffuse CK7+) vs. oncocytoma (rare sprinkled cells expressing CK7)

What specific CK7 staining patterns are diagnostically significant in challenging tumor types?

Certain CK7 staining patterns hold particular diagnostic significance:

Renal Tumors:

• Diffuse CK7 positivity supports chromophobe renal cell carcinoma

• Scattered or focal CK7 positivity may be seen in oncocytoma

• Clear cell papillary renal cell carcinoma typically shows strong and diffuse CK7 positivity

• Integration of CK7 with CD117 and Claudin-7 (the "three 7 markers") enhances diagnostic specificity for chromophobe RCC

Primary Extramammary Paget Disease:

• CK7 positivity serves as an excellent screening marker for primary extramammary Paget disease

• This distinguishes it from melanoma in situ (SOX10+), squamous cell carcinoma (p63+), and pagetoid urothelial intraepithelial neoplasia (uroplakin+)

Merkel Cell Carcinoma:

• Typically CK20+ with a characteristic perinuclear punctate or dot-like pattern

• Usually CK7 negative (only 23% CK7+), helping to distinguish it from other small round blue cell tumors

Cervical Lesions:

• CK7 expression in cervical low-grade squamous intraepithelial lesions may predict higher risk of progression to high-grade lesions

What are the emerging research applications of CK7 antibody beyond traditional diagnostics?

Beyond standard diagnostic applications, CK7 antibody is finding new applications in cutting-edge research:

Cancer Stem Cell Identification:
Researchers are investigating the relationship between CK7 expression and cancer stem cell properties in various tumor types. Variable CK7 expression patterns may correlate with different stemness properties and therapeutic resistance mechanisms.

Liquid Biopsy Development:
CK7 antibody is being evaluated for circulating tumor cell (CTC) detection and characterization in blood samples. This application could enable less invasive monitoring of disease progression and treatment response.

Precision Medicine Approaches:
Integration of CK7 into comprehensive biomarker panels for precision medicine applications represents a significant research direction, enabling more tailored treatment approaches based on molecular tumor characteristics.

What technical limitations must researchers consider when working with CK7 antibody?

Despite its utility, researchers should be aware of several technical considerations:

Species Cross-Reactivity:
Most CK7 antibody clones lack murine reactivity, which complicates xenograft studies. Careful evaluation of species cross-reactivity is essential when designing animal model research.

Epitope Accessibility:
Fixation conditions can significantly impact epitope accessibility. Prolonged formalin fixation may reduce CK7 immunoreactivity, requiring optimized antigen retrieval protocols.

Clone Selection Impact:
Different antibody clones may have variable sensitivities to different CK7 epitopes. For example, the rabbit monoclonal BC1 clone shows superior staining intensity compared to the mouse monoclonal OV-TL 12/30 clone in certain applications .

Interpretation Standardization:
The lack of standardized positivity thresholds across laboratories (e.g., >1% vs. >10% positivity) creates challenges for comparative studies and meta-analyses. Researchers should clearly define and justify their positivity criteria.