MAP3K7 (Ab-187) Antibody

What is MAP3K7 and why is it significant in cell signaling research?

MAP3K7/TAK1 (EC 2.7.11.25) is a serine/threonine kinase that plays a crucial role in various cellular signaling pathways, including those involving immune responses, inflammation, and tissue repair . It serves as a key mediator in the MAPK signaling cascade, integrating signals from cytokines like TGF-β and activating downstream effectors. Research has shown its involvement in multiple physiological and pathological processes, making it a significant target for investigation in disease mechanisms. In recent studies, MAP3K7 loss has been associated with enhanced androgen signaling and progression in prostate cancer, highlighting its potential tumor suppressor role in specific contexts .

What specifically does the MAP3K7 (Ab-187) antibody recognize?

The MAP3K7 (Ab-187) antibody specifically recognizes the region surrounding the threonine 187 phosphorylation site in human MAP3K7 protein. This antibody is generated using a synthesized non-phosphopeptide derived from human MAP3K7 with the amino acid sequence around the phosphorylation site of threonine 187 (H-M-T(p)-N-N) . This specificity allows researchers to detect the non-phosphorylated form of MAP3K7 at this critical regulatory site.

How does MAP3K7 (Ab-187) antibody differ from phospho-specific MAP3K7 antibodies?

Unlike phospho-specific antibodies that recognize only the phosphorylated form of MAP3K7 (such as phospho-MAP3K7-T187, phospho-MAP3K7-S412, or phospho-MAP3K7-S439) , the MAP3K7 (Ab-187) antibody detects the non-phosphorylated threonine 187 region. This distinction is crucial for experimental design as it allows researchers to measure total MAP3K7 protein levels independently of its phosphorylation status, providing complementary information to phospho-specific detection. When used alongside phospho-specific antibodies, researchers can calculate the ratio of phosphorylated to total protein to assess activation status.

What are the validated applications for MAP3K7 (Ab-187) antibody?

Based on the technical specifications from multiple suppliers, the MAP3K7 (Ab-187) antibody has been validated for several research applications:

These applications enable comprehensive investigation of MAP3K7 expression, localization, and function across diverse experimental systems .

How should I optimize Western blot protocols for MAP3K7 (Ab-187) antibody?

For optimal Western blot results with MAP3K7 (Ab-187) antibody:

• Sample preparation: Use standard protein isolation protocols with protease inhibitors to prevent degradation

• Loading: 20-40 μg of total protein per lane is typically sufficient

• Transfer: Standard wet or semi-dry transfer to PVDF membrane

• Blocking: 5% non-fat milk or BSA in TBST for 1 hour at room temperature

• Primary antibody: Dilute MAP3K7 (Ab-187) antibody at 1:500-1:2500 in blocking buffer and incubate overnight at 4°C

• Secondary antibody: Anti-rabbit IgG conjugated with HRP or fluorescent tags

• Detection: Compatible with both chemiluminescence and fluorescence-based detection systems, such as LI-COR Odyssey imaging systems

For quantification, normalize MAP3K7 signals to loading controls such as β-actin, α-tubulin, or Lamin A/C for nuclear extracts .

What are key considerations for successful immunohistochemistry with MAP3K7 (Ab-187) antibody?

When performing immunohistochemistry with MAP3K7 (Ab-187) antibody:

• Tissue preparation: Both formalin-fixed paraffin-embedded (FFPE) and frozen sections are compatible

• Antigen retrieval: Heat-induced epitope retrieval in citrate buffer (pH 6.0) is typically effective

• Blocking: 5-10% normal serum from the same species as the secondary antibody

• Primary antibody: Use at 1:50-1:80 dilution and incubate overnight at 4°C

• Detection system: Standard avidin-biotin or polymer-based detection systems

• Controls: Include positive controls (tissues known to express MAP3K7) and negative controls (primary antibody omitted)

When evaluating prostate cancer specimens, careful correlation of MAP3K7 expression with clinical parameters may provide valuable prognostic information, as low MAP3K7 expression has been identified as a strong independent predictor for biochemical recurrence .

What critical controls should I include when using MAP3K7 (Ab-187) antibody?

For rigorous experimental design:

• Positive control: Include cell lines or tissues with known MAP3K7 expression (e.g., NIH/3T3, K-562, SKOV-3, A375, A431 cells)

• Negative control: Use samples with minimal MAP3K7 expression or employ antibody diluent without primary antibody

• Knockdown/knockout control: When available, include MAP3K7-depleted samples to confirm specificity

• Loading/processing controls: Use housekeeping proteins (β-actin, GAPDH) for Western blots or standardized staining protocols for IHC

• Cross-validation: Consider validating key findings with a second MAP3K7 antibody recognizing a different epitope

These controls help establish specificity and reliability, particularly when investigating MAP3K7 in novel experimental contexts.

What are common troubleshooting issues when working with MAP3K7 (Ab-187) antibody?

When investigating MAP3K7 in cancer models, be aware that expression patterns may vary significantly between different tumor types and stages, requiring careful optimization of detection conditions.

How can I verify the specificity of MAP3K7 (Ab-187) antibody in my experimental system?

To verify antibody specificity:

• siRNA/shRNA knockdown: Compare signal in MAP3K7-depleted versus control samples

• Recombinant protein competition: Pre-incubate antibody with excess purified MAP3K7 protein before application

• Peptide blocking: Block with the immunizing peptide (179-194aa region of human MAP3K7)

• Orthogonal validation: Compare results with alternative detection methods (e.g., mass spectrometry)

• Cross-species validation: Test in samples from different species to confirm expected reactivity patterns with human, mouse, and rat samples

These validation approaches are particularly important when studying novel pathways or disease models where MAP3K7 function may be altered.

How can MAP3K7 (Ab-187) antibody be used to investigate signaling pathway crosstalk?

The MAP3K7 (Ab-187) antibody can be employed in several advanced approaches to study signaling pathway interactions:

• Co-immunoprecipitation: Use the antibody to pull down MAP3K7 complexes and identify interacting partners through mass spectrometry or Western blotting

• Proximity ligation assays: Combine with antibodies against potential interaction partners to visualize protein-protein interactions in situ

• Phosphorylation dynamics: Use in combination with phospho-specific antibodies (such as phospho-MAP3K7-T187, phospho-MAP3K7-S412, or phospho-MAP3K7-S439) to track activation patterns following various stimuli

• Chromatin immunoprecipitation: When studying nuclear translocation of MAP3K7, ChIP-seq can identify genomic targets

These approaches can reveal important insights into how MAP3K7 integrates signals from multiple upstream pathways and influences downstream effectors in different cellular contexts.

What can MAP3K7 (Ab-187) antibody reveal about disease mechanisms in cancer research?

In cancer research, MAP3K7 (Ab-187) antibody has proven valuable for understanding disease mechanisms:

• Expression analysis: Quantify MAP3K7 levels across tumor stages and types to identify correlations with disease progression

• Biomarker potential: Evaluate MAP3K7 as a prognostic indicator, particularly in prostate cancer where low MAP3K7 expression has been associated with biochemical recurrence

• Therapeutic response: Monitor changes in MAP3K7 expression or localization in response to treatments

• Functional studies: Combine with manipulation of MAP3K7 levels to understand its role in cancer cell survival, proliferation, and resistance mechanisms

Recent research has demonstrated that MAP3K7 loss can drive enhanced androgen signaling and increased AR-v7 expression in prostate cancer, contributing to enzalutamide resistance . This highlights the potential of MAP3K7 as both a biomarker and therapeutic target.

How should I quantify and normalize MAP3K7 expression data?

For robust quantification of MAP3K7 expression:

• Western blot densitometry:

  • Use linear range of detection (validate with dilution series)

  • Normalize to appropriate loading controls (β-actin, GAPDH)

  • Use software like Image Studio (LI-COR) for accurate quantification

• Immunohistochemistry scoring:

  • Implement standardized scoring systems (H-score, Allred, etc.)

  • Consider both staining intensity and percentage of positive cells

  • Employ digital pathology tools when available for objective quantification

• Statistical analysis:

  • Use appropriate statistical tests based on data distribution

  • Include sufficient biological and technical replicates

  • Consider multiple comparison corrections when analyzing across different conditions or timepoints

When studying MAP3K7 in cancer contexts, correlate expression data with clinical parameters and other molecular markers to enhance biological interpretation.

How can I address contradictory results when studying MAP3K7 across different experimental models?

When encountering contradictory MAP3K7 results:

• Consider cell/tissue specificity: MAP3K7 functions can vary dramatically between different cell types and tissues

• Evaluate activation status: Phosphorylation at different sites may result in distinct functional outcomes

• Assess experimental conditions: Growth factors, stress conditions, and cell density can influence MAP3K7 signaling

• Examine protein complexes: MAP3K7 functions within multi-protein complexes that may vary between models

• Check for genetic alterations: Mutations or deletions may be present in some cell lines or patient samples

In prostate cancer research, for example, the finding that MAP3K7 loss is the strongest independent predictor for biochemical recurrence over other tested clinicopathologic factors including CHD1 expression highlights the importance of considering MAP3K7 in the context of other genetic alterations.

What methodological considerations are important when using MAP3K7 (Ab-187) antibody in multiparameter analyses?

For multiparameter analyses incorporating MAP3K7 (Ab-187) antibody:

• Multiplexed immunofluorescence:

  • Ensure antibody compatibility with multiplexing reagents

  • Establish appropriate antibody order in sequential staining protocols

  • Validate signal specificity in the multiplexed context

• Flow cytometry applications:

  • Optimize fixation and permeabilization for intracellular MAP3K7 detection

  • Establish compensation controls when using multiple fluorochromes

  • Validate with appropriate positive and negative controls

• Single-cell analysis:

  • Consider cell-to-cell variability in MAP3K7 expression and activation

  • Correlate with other markers to identify distinct cellular subpopulations

  • Integrate with transcriptomic or proteomic data for comprehensive analysis

When designing such experiments, careful attention to antibody dilution, incubation conditions, and signal amplification methods is essential for generating reliable and reproducible results.