sst6 Antibody

What is STAT6 and what biological role does it play?

STAT6 (Signal Transducer and Activator of Transcription 6) is a member of the STAT family of transcription factors that plays a central role in mediating IL-4 and IL-13 signaling. In response to cytokines and growth factors, STAT family members are phosphorylated by receptor-associated kinases, forming homo- or heterodimers that translocate to the cell nucleus where they function as transcription activators . STAT6 is crucial for exerting IL4-mediated biological responses and induces the expression of BCL2L1/BCL-X(L), which is responsible for the anti-apoptotic activity of IL4 . Additionally, STAT6 drives Th2 immune responses and allergic inflammation, making it implicated in asthma and atopic diseases .

What types of STAT6 antibodies are available for research?

STAT6 antibodies are available in several formats to accommodate various research applications:

The selection of antibody type should be based on the specific research application, with rabbit monoclonal antibodies often providing excellent specificity for immunohistochemistry applications .

How should STAT6 antibodies be stored and handled to maintain activity?

While specific storage conditions may vary by manufacturer, generally STAT6 antibodies should be stored according to these guidelines to maintain optimal activity:

• For long-term storage, keep antibodies at -20°C to prevent degradation

• Avoid repeated freeze-thaw cycles by aliquoting antibodies into single-use volumes

• For short-term storage (up to 1 month), antibodies can typically be kept at 4°C

• Follow manufacturer-specific recommendations for buffer conditions and stabilizers

• Protect conjugated antibodies (especially fluorescent conjugates) from light exposure

Proper storage and handling are essential for maintaining antibody performance and experimental reproducibility .

What applications are STAT6 antibodies suitable for?

STAT6 antibodies can be employed in various laboratory applications depending on the research question:

The choice of application should be guided by the specific research objectives and the validated performance of the antibody for each technique .

How can STAT6 antibodies be used for the differential diagnosis of soft tissue tumors?

STAT6 immunohistochemistry has emerged as a powerful diagnostic tool for Solitary Fibrous Tumors (SFT). Studies have demonstrated that nuclear STAT6 staining is present in virtually all SFT cases (sensitivity 100%), regardless of histology, anatomical site, or CD34 status. The underlying molecular basis for this diagnostic utility is the presence of recurrent NAB2-STAT6 gene fusions specifically in SFTs .

For optimal diagnostic application:

• Use rabbit monoclonal STAT6 antibody at appropriate dilution (typically 1:100)

• Evaluate only nuclear staining as positive (cytoplasmic staining should be disregarded)

• Consider staining distribution scoring: 0 (no staining), 1+ (1-25%), 2+ (26-50%), 3+ (>50%)

• Assess staining intensity as weak, moderate, or strong

Most SFTs show 3+ and strong staining patterns. Importantly, other morphologically similar soft tissue tumors including cellular angiofibroma, myofibroblastoma, spindle cell lipoma, dermatofibrosarcoma protuberans, and various sarcomas consistently show negative results for nuclear STAT6 staining (specificity 100%) . This makes STAT6 immunohistochemistry an invaluable tool in the differential diagnosis of soft tissue tumors.

What are the potential pitfalls in using STAT6 as a diagnostic marker?

Despite the high sensitivity and specificity of STAT6 as a diagnostic marker for Solitary Fibrous Tumors, researchers should be aware of potential pitfalls:

• STAT6 amplification occurs in a subset of dedifferentiated Liposarcomas, resulting in STAT6 protein expression detectable by immunohistochemistry. This represents a potential diagnostic pitfall in the differential diagnosis of dedifferentiated Liposarcoma and Solitary Fibrous Tumor .

• Weak STAT6 staining has been reported in a small number of deep Fibrous Histiocytomas and dedifferentiated Liposarcomas . Therefore:

  • Evaluate staining intensity and pattern carefully

  • Consider correlating with morphological features

  • Use additional markers when appropriate for challenging cases

  • Be cautious of interpretation when only weak staining is observed

• Technical factors affecting staining quality:

  • Fixation time and conditions

  • Antigen retrieval methods

  • Antibody clone and dilution

  • Detection systems

Understanding these limitations is crucial for accurate interpretation of STAT6 immunohistochemistry in diagnostic pathology .

How can phospho-specific STAT6 antibodies be utilized to study signaling pathways?

Phospho-specific STAT6 antibodies provide valuable tools for studying the activation status of STAT6 signaling pathways. These antibodies target specific phosphorylation sites, particularly Tyr641 and Thr645, which are critical for STAT6 activation and function .

Methodological approaches for phospho-STAT6 research:

• Stimulation experiments:

  • Treat cells with IL-4 or IL-13 to induce STAT6 phosphorylation

  • Harvest cells at different time points to evaluate phosphorylation kinetics

  • Compare phospho-STAT6 levels with total STAT6 expression

• Inhibitor studies:

  • Use JAK inhibitors to block STAT6 phosphorylation

  • Evaluate dose-dependent effects on signaling pathway

  • Monitor downstream effects on target gene expression

• Mutation analysis:

  • Compare wild-type and mutant STAT6 phosphorylation patterns

  • Assess the impact of specific mutations on signaling cascade

  • Correlate with functional outcomes

• Multiplex signaling analysis:

  • Combine phospho-STAT6 detection with other pathway components

  • Map cross-talk between STAT6 and related signaling pathways

  • Develop integrated models of cellular signaling networks

These approaches enable detailed mechanistic studies of STAT6-dependent processes in allergy, inflammation, and cancer research .

What controls should be included when validating STAT6 antibodies for experimental use?

Proper validation of STAT6 antibodies is essential for generating reliable and reproducible research data. A comprehensive validation approach should include multiple controls:

• Positive controls:

  • Cell lines with known STAT6 expression (e.g., lymphocytes stimulated with IL-4)

  • Tissue samples with established STAT6 expression patterns

  • Recombinant STAT6 protein (for Western blot)

  • Known STAT6-positive tumors (e.g., Solitary Fibrous Tumors for IHC)

• Negative controls:

  • STAT6 knockout or knockdown samples

  • Cell lines lacking STAT6 expression

  • Tissue samples from STAT6-deficient models

  • Pre-absorption with immunizing peptide

• Specificity controls:

  • Cross-reactivity testing with other STAT family members

  • Epitope mapping to confirm binding region

  • Comparison of multiple antibody clones targeting different epitopes

  • Validation across multiple applications (WB, IHC, IF)

• Technical controls:

  • Secondary antibody-only controls to assess background

  • Isotype controls to evaluate non-specific binding

  • Titration experiments to determine optimal concentrations

  • Reproducibility testing across different lots

Documentation of these validation steps enhances confidence in experimental results and facilitates troubleshooting when inconsistencies arise .

What are common causes of false positive or false negative results with STAT6 antibodies?

Understanding potential sources of false results is crucial for accurate interpretation:

Causes of false positive results:

• Cross-reactivity with other STAT family members due to sequence homology

• Non-specific binding, particularly with polyclonal antibodies

• Excessive antibody concentration leading to background staining

• Inadequate blocking of endogenous peroxidases or biotin

• Inappropriate detection system amplification

Causes of false negative results:

• Insufficient antigen retrieval, especially for formalin-fixed tissues

• Antibody degradation due to improper storage

• Epitope masking during tissue processing

• Suboptimal incubation conditions (time, temperature, concentration)

• Use of antibodies not validated for specific applications

Mitigation strategies:

• Optimize protocols for each specific application

• Include appropriate positive and negative controls

• Validate antibodies using multiple methods

• Consider using monoclonal antibodies for higher specificity

• Follow manufacturer's recommendations for sample preparation

How can researchers optimize STAT6 antibody immunohistochemistry protocols?

Optimization of immunohistochemistry protocols for STAT6 detection involves several critical considerations:

• Fixation and tissue processing:

  • Standardize fixation time (typically 24-48 hours in 10% neutral buffered formalin)

  • Ensure complete tissue processing and proper paraffin embedding

  • Use freshly cut sections (4-5 μm thickness optimal)

• Antigen retrieval:

  • Heat-induced epitope retrieval (HIER) methods often yield best results

  • Test multiple pH conditions (citrate buffer pH 6.0 vs. EDTA buffer pH 9.0)

  • Optimize retrieval duration and temperature

• Antibody selection and dilution:

  • Rabbit monoclonal antibodies often provide superior results for STAT6

  • Perform titration experiments to determine optimal concentration

  • Consider the clone specificity (e.g., EP325 clone has demonstrated high sensitivity)

• Detection system:

  • Polymer-based detection systems typically offer improved sensitivity

  • Minimize background by optimizing incubation times

  • Consider signal amplification for low-abundance targets

• Evaluation and interpretation:

  • Focus on nuclear staining patterns for diagnostic applications

  • Use standardized scoring systems for intensity and distribution

  • Document both positive and negative staining areas

When optimizing for Solitary Fibrous Tumor diagnosis, nuclear staining should be the primary focus, as it represents the translocation of activated STAT6 to the nucleus .

What strategies can be employed to resolve discrepant STAT6 antibody results across different applications?

When facing discrepant results with STAT6 antibodies across different applications, systematic troubleshooting is essential:

• Antibody-specific factors:

  • Determine if the antibody is validated for all attempted applications

  • Consider epitope accessibility in different applications (linear vs. conformational)

  • Test alternative antibody clones targeting different epitopes

  • Verify antibody lot consistency and consider lot-to-lot variation

• Sample preparation considerations:

  • Native vs. denatured protein detection requirements

  • Fixation effects on epitope availability

  • Buffer compatibility with specific applications

  • Protein modification status (phosphorylation, glycosylation)

• Technical approach:

  • Create a validation matrix across applications

  • Use orthogonal methods to confirm findings

  • Implement positive and negative controls for each application

  • Standardize protocols between experiments

• Reconciliation strategies:

  • Identify pattern-based discrepancies to determine systematic issues

  • Consider biological variables (cell type, stimulation status)

  • Evaluate technical variables (equipment, reagents, operators)

  • Document comprehensive method details for meaningful comparisons

Through systematic evaluation of these factors, researchers can reconcile discrepant results and develop more robust protocols for consistent STAT6 detection .

How are STAT6 antibodies being used in coronavirus and immune response research?

STAT6 antibodies are being incorporated into research examining immune responses to viral infections, including coronaviruses:

• Antibody screening approaches:
  Recent studies have developed highly specific and sensitive approaches for measuring antibodies against SARS-CoV-2 for population-scale immune surveillance. These methods often define antibody positivity through dual-positive responses against multiple viral proteins (e.g., receptor-binding domain and nucleocapsid proteins) .

• STAT6 pathway in viral immunity:

  • Investigation of STAT6 signaling in type 2 immune responses during viral infection

  • Examination of IL-4/IL-13 axis modulation during coronavirus infection

  • Analysis of STAT6 activation in immune cells responding to viral challenge

• Epitope scaffold design:
  Advanced immunogen engineering approaches incorporate epitope grafting to design scaffolds that strongly interact with broadly cross-reactive antibodies. While not directly related to STAT6 antibodies, these approaches represent cutting-edge methodologies in antibody research that may be applicable to studying immune regulation through the STAT6 pathway .

• Therapeutic implications:
  Understanding STAT6 signaling in the context of viral infections and immune responses may provide insights into novel therapeutic approaches, particularly for conditions involving dysregulated Th2 responses .

This emerging research area highlights the interconnection between STAT6 signaling pathways and broader immune responses to viral pathogens.

What are the latest advancements in STAT6 antibody technology for cancer research?

Recent innovations in STAT6 antibody technology are expanding capabilities in cancer research:

• Diagnostic applications:
  The discovery that nuclear STAT6 expression (detected by immunohistochemistry) serves as a highly sensitive and specific marker for Solitary Fibrous Tumors has revolutionized the diagnosis of these neoplasms. This finding is based on the identification of recurrent NAB2-STAT6 gene fusions located at chromosomal region 12q13 in SFTs .

• Prognostic significance:
  Researchers are investigating correlations between STAT6 expression patterns, activation status, and patient outcomes in various malignancies, including:

  • Hematological malignancies

  • Solid tumors with STAT6 pathway alterations

  • Tumors with immune microenvironment modulation

• Therapeutic target validation:
  STAT6 antibodies are increasingly used to validate this pathway as a potential therapeutic target in:

  • Tumors with STAT6 genomic alterations

  • Malignancies with IL-4/IL-13 driven progression

  • Cancer types with immunosuppressive tumor microenvironments

• Technical innovations:

  • Development of phospho-specific antibodies for detailed signaling studies

  • Multiplex immunohistochemistry to correlate STAT6 with other markers

  • Enhanced sensitivity detection systems for low-abundance detection

These advancements demonstrate the evolving role of STAT6 antibodies in cancer research, from diagnostic biomarkers to tools for understanding complex tumor biology .

How can researchers design experiments to investigate STAT6 phosphorylation dynamics in different cell types?

Designing experiments to investigate STAT6 phosphorylation dynamics requires careful consideration of multiple factors:

• Cell system selection:

  • Primary cells vs. cell lines (consider physiological relevance)

  • Relevant tissue types (lymphocytes, macrophages, epithelial cells)

  • Genetic modification options (CRISPR, RNAi) for pathway component manipulation

• Stimulation paradigms:

  • Dose-response curves for IL-4 and IL-13

  • Time-course experiments (ranging from minutes to hours)

  • Combinatorial stimulation with other cytokines to assess pathway cross-talk

  • Physiological vs. pathological concentration ranges

• Detection methods:

  • Phospho-specific STAT6 antibodies (targeting Tyr641 or Thr645)

  • Western blotting for bulk analysis

  • Flow cytometry for single-cell resolution

  • Immunofluorescence for spatial information

  • Phospho-proteomics for comprehensive pathway analysis

• Experimental design:

  • Include both total STAT6 and phospho-STAT6 measurements

  • Monitor nuclear translocation (fractionation or imaging)

  • Assess downstream gene activation (qPCR, reporter assays)

  • Incorporate pathway inhibitors at different levels

• Data analysis approaches:

  • Quantification of phosphorylation kinetics

  • Correlation with functional outcomes

  • Mathematical modeling of signaling dynamics

  • Integration with other pathway components

This experimental framework enables detailed characterization of STAT6 phosphorylation dynamics across different cellular contexts and stimulation conditions .