Phospho-MAP3K1 (Thr1402) Antibody

What is Phospho-MAP3K1 (Thr1402) Antibody and what cellular processes does it help investigate?

Phospho-MAP3K1 (Thr1402) Antibody is a rabbit polyclonal antibody specifically designed to detect endogenous levels of MAP3K1 protein only when phosphorylated at threonine 1402 . MAP3K1 (also known as MEKK1) functions as a critical component of protein kinase signal transduction cascades, primarily activating the ERK and JNK kinase pathways through phosphorylation of MAP2K1 and MAP2K4 . Additionally, MAP3K1 activates CHUK and IKBKB, the central protein kinases of the NF-kappa-B pathway . This antibody enables researchers to investigate cellular signaling mechanisms related to stress responses, inflammation, apoptosis, and cell proliferation where MAP3K1 phosphorylation plays a significant role.

What are the validated experimental applications for Phospho-MAP3K1 (Thr1402) Antibody?

The Phospho-MAP3K1 (Thr1402) Antibody has been validated for multiple research applications:

This antibody has been specifically validated in Western blot analysis of lysates from Jurkat cells and NIH-3T3 cells, as well as in immunohistochemistry analysis of paraffin-embedded human brain tissue . The antibody demonstrates high specificity, as shown by blocking experiments with the phospho-peptide that eliminate the detection signal .

How does the specificity of this antibody compare to other MAP3K1 antibodies?

The Phospho-MAP3K1 (Thr1402) Antibody is highly specific for the phosphorylated form of MAP3K1 at threonine 1402, distinguishing it from general MAP3K1 antibodies that detect the protein regardless of phosphorylation status . This specificity is achieved through a rigorous purification process. The antibodies are first produced by immunizing rabbits with synthetic phosphopeptide and KLH conjugates . They are then purified by affinity-chromatography using epitope-specific phosphopeptide . Importantly, non-phospho specific antibodies are removed by chromatography using non-phosphopeptide . This dual purification approach results in an antibody that exclusively recognizes MAP3K1 when phosphorylated at Thr1402, enabling researchers to specifically track this post-translational modification in experimental contexts.

What are the optimal sample preparation conditions for detecting phospho-MAP3K1 in Western blotting?

For optimal detection of phosphorylated MAP3K1 in Western blotting, researchers should follow these methodological steps:

• Cell lysis should be performed using a phosphatase inhibitor-containing buffer to preserve the phosphorylation state of MAP3K1 at Thr1402. Common inhibitors include sodium orthovanadate, sodium fluoride, and phosphatase inhibitor cocktails.

• Sample preparation should include denaturation in Laemmli buffer at 95°C for 5 minutes. Given MAP3K1's large size (161kDa), use a lower percentage (6-8%) SDS-PAGE gel for better resolution .

• For protein transfer, a wet transfer system is recommended, running at lower voltage (30V) overnight at 4°C to ensure complete transfer of the high molecular weight protein.

• Blocking should be performed with 5% BSA in TBST (not milk, which contains phosphatases that could reduce signal).

• The antibody should be diluted 1:500-1:1000 in 5% BSA/TBST and incubated overnight at 4°C for optimal binding .

• Particular attention should be paid to including positive controls (e.g., Jurkat or NIH-3T3 cell lysates) with known MAP3K1 phosphorylation, as demonstrated in validation studies .

How should tissue samples be prepared for immunohistochemistry with Phospho-MAP3K1 (Thr1402) Antibody?

For immunohistochemistry applications, tissue sample preparation requires specific considerations:

• Fixation should be performed with 10% neutral-buffered formalin for 24-48 hours, followed by paraffin embedding using standard protocols.

• Tissue sections should be cut at 4-6μm thickness and mounted on positively charged slides.

• Antigen retrieval is critical for phospho-epitopes and should be performed using citrate buffer (pH 6.0) or EDTA buffer (pH 9.0) in a pressure cooker or microwave.

• Prior to antibody incubation, endogenous peroxidase activity should be quenched with 3% hydrogen peroxide, and non-specific binding blocked with 5% normal goat serum.

• The antibody should be applied at a dilution of 1:50-1:100 and incubated overnight at 4°C in a humidified chamber .

• Detection should use a polymer-based detection system rather than biotin-avidin systems to avoid background issues.

• Always include a negative control (omitting primary antibody) and a competing phospho-peptide control to confirm specificity, as demonstrated in the validation studies with human brain tissue .

What controls should be included to validate Phospho-MAP3K1 (Thr1402) antibody specificity in experiments?

To ensure experimental rigor when using Phospho-MAP3K1 (Thr1402) Antibody, the following controls should be incorporated:

• Positive Control: Include lysates from cells known to express phosphorylated MAP3K1, such as Jurkat cells or NIH-3T3 cells as demonstrated in validation studies .

• Negative Control: Use samples from cells where MAP3K1 phosphorylation has been inhibited or cells known not to express MAP3K1.

• Peptide Competition Assay: Perform parallel experiments where the antibody is pre-incubated with the immunizing phospho-peptide, which should abolish specific staining. This control is particularly important and has been validated in both Western blot and immunohistochemistry applications .

• Phosphatase Treatment Control: Treat a duplicate sample with lambda phosphatase to remove phosphorylation, which should eliminate the signal when using the phospho-specific antibody.

• Loading Control: For Western blotting, include an antibody against total MAP3K1 or a housekeeping protein on a separate blot prepared with identical samples.

• Secondary Antibody Control: Include a sample incubated only with secondary antibody to identify potential non-specific binding.

These controls collectively ensure that the observed signals are genuinely attributable to Phospho-MAP3K1 (Thr1402) rather than experimental artifacts.

How can researchers address weak or absent signals when using Phospho-MAP3K1 (Thr1402) Antibody?

When encountering weak or absent signals with Phospho-MAP3K1 (Thr1402) Antibody, researchers should systematically address potential issues:

• Phosphorylation Preservation: Ensure immediate sample processing with phosphatase inhibitors (sodium orthovanadate, sodium fluoride, etc.) in all buffers. Phosphorylation can be extremely labile, particularly during extended handling.

• Antibody Concentration: If signal is weak, try increasing the antibody concentration from the recommended 1:1000 to 1:500 for Western blotting or from 1:100 to 1:50 for immunohistochemistry .

• Incubation Conditions: Extend primary antibody incubation time to overnight at 4°C and ensure gentle agitation for uniform antibody access.

• Protein Loading: For Western blotting, increase protein loading to 50-80μg total protein per lane, as phosphorylated MAP3K1 may be present at low levels.

• Detection System Enhancement: Use high-sensitivity chemiluminescent substrates for Western blotting or signal amplification systems (TSA) for immunohistochemistry.

• Cell Stimulation: Consider pre-treating cells with stimulants known to increase MAP3K1 phosphorylation at Thr1402 before sample collection.

• Storage Conditions: Verify the antibody has been stored properly at -20°C or -80°C and has not undergone multiple freeze-thaw cycles, which can degrade activity .

• Antibody Lot: Different lots may have variation in sensitivity; consider validating with a positive control known to work with the specific antibody lot.

What are common causes of non-specific binding and how can they be minimized?

Non-specific binding can significantly impact experimental results when using Phospho-MAP3K1 (Thr1402) Antibody. Here are methodological approaches to address this issue:

• Blocking Optimization: Increase blocking time (2-3 hours at room temperature) and test different blocking agents. For phospho-antibodies, 5% BSA in TBST is typically superior to milk-based blockers, which contain phosphatases.

• Antibody Dilution: Further dilute the antibody if background is excessive. While recommendations suggest 1:500-1:1000 for Western blotting, testing a dilution series (1:500, 1:1000, 1:2000) can help identify optimal conditions .

• Washing Protocols: Implement more stringent washing steps (5-6 washes of 10 minutes each) with 0.1% Tween-20 in TBS after both primary and secondary antibody incubations.

• Secondary Antibody Cross-Reactivity: Use secondary antibodies specifically adsorbed against other species to reduce cross-reactivity.

• Tissue Autofluorescence: For immunofluorescence applications, pretreat sections with sodium borohydride or commercial autofluorescence quenching reagents.

• Endogenous Peroxidase/Biotin: For IHC applications, ensure thorough quenching of endogenous peroxidase with 3% H₂O₂ and use biotin-free detection systems if endogenous biotin is problematic.

• Antibody Purification: The Phospho-MAP3K1 (Thr1402) Antibody undergoes dual purification, including removal of non-phospho specific antibodies by chromatography using non-phosphopeptide . Ensure the antibody used has undergone this rigorous purification process.

• Competing Peptide Controls: Always run a competing phospho-peptide control to distinguish between specific and non-specific binding, as demonstrated in the validation studies .

How should researchers modify protocols when working with different sample types (cell lines vs. primary tissues)?

Protocol modifications are essential when transitioning between different sample types:

For Cell Lines:

• Lysis Buffer Composition: Standard RIPA or NP-40 buffers supplemented with phosphatase inhibitors are typically sufficient for cell line samples.

• Stimulation Protocols: Consider appropriate stimulation to enhance phosphorylation. For example, stress inducers or growth factors can be used to activate MAP3K1 signaling prior to sample collection.

• Antibody Dilution: Start with the recommended dilution of 1:500-1:1000 for Western blotting .

• Signal Detection: Standard chemiluminescence detection systems are usually adequate.

For Primary Tissues:

• Tissue Handling: Minimize warm ischemia time and flash-freeze samples immediately after collection to preserve phosphorylation states.

• Extraction Protocol: Use more stringent extraction methods, such as tissue homogenization in RIPA buffer with increased detergent concentrations and mechanical disruption.

• Antibody Concentration: Often requires higher antibody concentrations (1:50-1:100 for IHC) compared to cell lines .

• Antigen Retrieval: More aggressive antigen retrieval methods are typically necessary, such as pressure cooking in citrate or EDTA buffer.

• Signal Amplification: Consider tyramide signal amplification (TSA) or other amplification systems for IHC/IF to enhance detection sensitivity.

• Background Reduction: Additional blocking steps with animal serum matching the host of the secondary antibody may be necessary to reduce non-specific binding in complex tissue samples.

• Validation: More extensive validation is required, including comparison with appropriate positive control tissues and competing peptide controls as demonstrated in the human brain tissue validation .

How can Phospho-MAP3K1 (Thr1402) Antibody be incorporated into multiplexed phospho-protein analysis workflows?

Integrating Phospho-MAP3K1 (Thr1402) Antibody into multiplexed phospho-protein analysis requires strategic methodological approaches:

• Sequential Immunoblotting: For Western blot-based multiplexing, researchers can use:

  • Mild stripping buffers (glycine-SDS, pH 2.5) that remove antibodies while preserving proteins on the membrane

  • Sequential probing with Phospho-MAP3K1 (Thr1402) Antibody and then other phospho-specific antibodies targeting related pathway components

  • Careful documentation of membrane orientation and protein ladder positions between stripping cycles

• Multiplex Immunofluorescence:

  • Select primary antibodies from different host species (Phospho-MAP3K1 is rabbit-derived)

  • Use directly conjugated secondary antibodies with non-overlapping fluorophores

  • Apply tyramide signal amplification (TSA) for sequential staining with antibodies from the same species

  • Include single-color controls to confirm specificity and absence of bleed-through

• Bead-Based Multiplex Assays:

  • Adapt the antibody for conjugation to microspheres with unique spectral signatures

  • Optimize antibody:bead ratios to ensure sensitivity without cross-reactivity

  • Include phospho-peptide competition controls to verify specificity in the multiplex format

• Mass Cytometry (CyTOF):

  • Metal-conjugate the Phospho-MAP3K1 (Thr1402) Antibody using commercial conjugation kits

  • Titrate the metal-conjugated antibody to determine optimal concentration

  • Include isotype controls and blocking peptide controls

• Spatial Analysis:

  • Combine with digital spatial profiling or multiplexed ion beam imaging (MIBI) for spatial context

  • Validate signal specificity through phospho-peptide competition in spatial contexts

• Normalization Strategy:

  • Always include total MAP3K1 antibody in the multiplexed panel

  • Use housekeeping proteins or structural markers appropriate for the sample type

  • Apply computational methods to account for signal variations between markers

What approaches can be used to correlate MAP3K1 Thr1402 phosphorylation with downstream pathway activation?

To establish meaningful correlations between MAP3K1 Thr1402 phosphorylation and downstream signaling events, researchers should implement the following analytical strategies:

• Temporal Phosphorylation Profiling:

  • Conduct time-course experiments following stimulation

  • Use Phospho-MAP3K1 (Thr1402) Antibody in parallel with antibodies detecting phosphorylated forms of downstream targets (MAP2K1, MAP2K4, JNK, ERK, CHUK, and IKBKB)

  • Apply statistical methods such as cross-correlation analysis to determine time lags between phosphorylation events

• Pharmacological Intervention Studies:

  • Employ specific kinase inhibitors targeting MAP3K1 or upstream activators

  • Monitor changes in both Thr1402 phosphorylation and downstream pathway components

  • Use dose-response curves to establish quantitative relationships between inhibition of MAP3K1 phosphorylation and downstream effects

• Genetic Manipulation Approaches:

  • Generate phospho-mimetic (T1402D or T1402E) and phospho-dead (T1402A) MAP3K1 mutants

  • Express these constructs in appropriate cellular models

  • Use the Phospho-MAP3K1 (Thr1402) Antibody to confirm specificity (should not detect T1402A mutant)

  • Measure downstream pathway activation in each mutant condition

• Proximity Ligation Assays (PLA):

  • Combine Phospho-MAP3K1 (Thr1402) Antibody with antibodies against potential interaction partners

  • Visualize and quantify protein-protein interactions dependent on Thr1402 phosphorylation status

• Phospho-proteomic Integration:

  • Perform phospho-proteomic analysis following MAP3K1 activation or inhibition

  • Correlate Thr1402 phosphorylation levels (measured by Western blot with the antibody) with global phosphorylation changes

  • Apply pathway enrichment analysis to identify coordinated regulation patterns

• Single-Cell Analysis:

  • Adapt the Phospho-MAP3K1 (Thr1402) Antibody for flow cytometry or mass cytometry

  • Correlate phosphorylation at the single-cell level with markers of pathway activation

  • Identify cell subpopulations with distinct signaling states

• In situ Analysis:

  • Perform sequential IHC or multiplexed immunofluorescence on tissue sections

  • Colocalize Phospho-MAP3K1 (Thr1402) with markers of downstream pathway activation

  • Quantify spatial correlation using digital image analysis

How can researchers quantitatively assess the relationship between MAP3K1 Thr1402 phosphorylation and biological outcomes?

Establishing quantitative relationships between MAP3K1 Thr1402 phosphorylation and biological outcomes requires rigorous experimental design and data analysis:

• Quantitative Western Blot Analysis:

  • Generate standard curves using recombinant phosphorylated protein or synthesized phospho-peptides

  • Implement densitometric analysis with appropriate normalization to total protein or housekeeping proteins

  • Calculate the phospho-MAP3K1/total MAP3K1 ratio to account for expression level variations

  • Apply the antibody at the validated dilutions of 1:500-1:1000

• Dose-Response Experiments:

  • Expose cells to varying concentrations of stimulus known to activate MAP3K1

  • Quantify Thr1402 phosphorylation using the antibody across the dose range

  • Measure corresponding biological outcomes (e.g., cell proliferation, apoptosis, gene expression)

  • Apply mathematical modeling (Hill equation, logistic regression) to define the relationship

• Temporal Resolution Analysis:

  • Establish detailed time courses of Thr1402 phosphorylation following stimulation

  • Correlate with time courses of biological responses

  • Apply time-series analysis methods to establish causal relationships

  • Consider implementing pulse-chase approaches to determine phosphorylation dynamics

• Single-Cell Correlation Studies:

  • Adapt the Phospho-MAP3K1 (Thr1402) Antibody for immunofluorescence at the recommended dilution of 1:50-1:100

  • Combine with assays measuring biological outcomes at the single-cell level

  • Apply quantitative image analysis to correlate phosphorylation intensity with outcome measures

  • Use machine learning approaches to identify patterns in heterogeneous cell populations

• Genetic Approach:

  • Generate cell lines with varying expression levels of wild-type or mutant MAP3K1

  • Quantify Thr1402 phosphorylation across these cell lines

  • Measure corresponding biological outcomes

  • Perform regression analysis to establish quantitative relationships

• Systems Biology Integration:

  • Incorporate phosphorylation data into computational models of the signaling pathway

  • Validate model predictions using the antibody to measure phosphorylation under various conditions

  • Use sensitivity analysis to determine the influence of Thr1402 phosphorylation on model outputs

• Clinical Sample Analysis:

  • Apply the antibody in IHC analysis of patient samples at recommended dilutions (1:50-1:100)

  • Score phosphorylation levels using established pathology criteria

  • Correlate with clinical outcomes using appropriate statistical methods

  • Consider survival analysis (Kaplan-Meier, Cox regression) to relate phosphorylation to patient prognosis

What are the optimal storage and handling conditions to maintain antibody performance?

To preserve antibody activity and specificity, researchers should adhere to these storage and handling guidelines:

• Long-term Storage:

  • Store the antibody at -20°C or -80°C as recommended by all suppliers

  • Avoid repeated freeze-thaw cycles which can degrade antibody quality

  • Consider aliquoting the antibody upon receipt into single-use volumes

• Working Solution Preparation:

  • When preparing working dilutions, use high-quality, sterile buffer

  • For Phospho-MAP3K1 (Thr1402) Antibody, prepare dilutions in buffers containing 5% BSA rather than milk

  • Use freshly prepared dilutions whenever possible

• Handling Precautions:

  • Minimize exposure to room temperature

  • Avoid contamination by using sterile technique

  • Never vortex the antibody; mix by gentle inversion or flicking

• Buffer Composition:

  • The antibody is supplied in phosphate buffered saline (without Mg²⁺ and Ca²⁺), pH 7.4, with 150mM NaCl, 0.02% sodium azide, and 50% glycerol

  • Maintain similar conditions when diluting to preserve structural integrity

• Transport Conditions:

  • The antibody is typically shipped at 4°C

  • Upon receipt, immediately transfer to -20°C or -80°C for long-term storage

• Contamination Prevention:

  • Use sterile pipette tips for each handling

  • Avoid touching the inside of tubes containing antibody

  • Consider using antibody stabilizers if preparing stocks for extended use

• Quality Monitoring:

  • Test antibody performance periodically with positive controls

  • Monitor for signs of degradation such as precipitation, cloudiness, or diminished signal

  • Document lot numbers and performance to track potential lot-to-lot variations

How can researchers validate lot-to-lot consistency of Phospho-MAP3K1 (Thr1402) Antibody?

Ensuring consistent performance across different antibody lots is critical for experimental reproducibility:

• Reference Sample Banking:

  • Maintain frozen aliquots of well-characterized positive control samples (e.g., Jurkat cells or NIH-3T3 cells)

  • Use these reference samples to test each new lot under identical conditions

• Quantitative Comparison:

  • Perform side-by-side Western blot analysis with both old and new antibody lots

  • Quantify signal intensity and background levels using densitometry

  • Calculate signal-to-noise ratios to objectively compare performance

• Epitope Verification:

  • Conduct peptide competition assays with each new lot

  • Compare the degree of signal reduction when pre-incubated with the phospho-peptide

  • Ensure the peptide sequence around the phosphorylation site of threonine 1402 (T-G-A(p)-G-F) is recognized

• Cross-platform Validation:

  • Test new lots in multiple applications (WB, IHC, ELISA) if the antibody will be used across these methods

  • Verify that recommended dilutions (WB: 1:500-1:1000, IHC: 1:50-1:100) produce comparable results

• Specificity Assessment:

  • Test reactivity against phosphatase-treated samples

  • Verify absence of signal in negative control samples

  • Confirm consistent species cross-reactivity (Human, Mouse, Rat)

• Documentation System:

  • Maintain detailed records of lot numbers, dates received, and performance metrics

  • Document any deviations or adjustments needed for specific lots

  • Consider implementing a laboratory information management system (LIMS) for tracking

• Supplier Communication:

  • Request lot-specific validation data from suppliers

  • Inquire about changes in production protocols or purification methods

  • Report significant performance variations to the supplier for investigation

What emerging technologies might enhance the utility of Phospho-MAP3K1 (Thr1402) Antibody in future research?

Several emerging technologies show promise for expanding the applications of Phospho-MAP3K1 (Thr1402) Antibody:

• Advanced Proximity Labeling:

  • Conjugating the antibody to engineered peroxidases or biotin ligases to identify proteins in proximity to phosphorylated MAP3K1

  • This could reveal context-specific interaction partners dependent on Thr1402 phosphorylation status

• Super-Resolution Microscopy:

  • Adapting the antibody for STORM, PALM, or STED microscopy to visualize subcellular localization of phosphorylated MAP3K1 at nanometer resolution

  • This would provide insights into spatial organization of signaling complexes

• Live-Cell Phosphorylation Sensors:

  • Developing conformation-sensitive fluorescent proteins that can report on Thr1402 phosphorylation in real-time

  • These could be calibrated using the phospho-specific antibody in fixed cells

• Tissue Clearing Techniques:

  • Combining the antibody with CLARITY, iDISCO, or other clearing methods to enable 3D visualization of phosphorylation patterns in intact tissues

  • This would reveal tissue-level organization of MAP3K1 signaling

• Spatially Resolved Transcriptomics:

  • Integrating antibody-based phospho-protein detection with spatial transcriptomics to correlate Thr1402 phosphorylation with gene expression patterns

  • This would connect signaling events to transcriptional outcomes with spatial context

• Mass Spectrometry Immunoprecipitation:

  • Using the antibody for immunoprecipitation followed by mass spectrometry to identify post-translational modification patterns co-occurring with Thr1402 phosphorylation

  • This would provide a systems-level view of the phosphorylation state of MAP3K1

• Single-Molecule Pull-Down:

  • Applying the antibody in single-molecule pull-down assays to study individual MAP3K1 molecules and their phosphorylation states

  • This would reveal heterogeneity in phosphorylation patterns not detectable in bulk assays

• Cryo-Electron Microscopy:

  • Using the antibody to facilitate structural studies of phosphorylated MAP3K1 complexes

  • This could reveal conformational changes induced by Thr1402 phosphorylation

What are the most promising research areas where Phospho-MAP3K1 (Thr1402) Antibody could provide new biological insights?

The Phospho-MAP3K1 (Thr1402) Antibody holds particular promise for advancing several critical research domains:

• Cancer Signaling Networks:

  • Investigating the role of MAP3K1 Thr1402 phosphorylation in cancer progression and treatment resistance

  • The antibody could help identify patients likely to respond to kinase inhibitor therapies

  • Applications in both Western blotting (1:500-1:1000) and IHC (1:50-1:100) would be valuable for translational research

• Inflammatory Response Regulation:

  • Exploring how MAP3K1 phosphorylation status affects NF-κB pathway activation in inflammatory diseases

  • The antibody could help map phosphorylation dynamics during acute and chronic inflammation

  • This would leverage MAP3K1's known role in activating CHUK and IKBKB, the central protein kinases of the NF-kappa-B pathway

• Neuronal Stress Responses:

  • Examining the role of MAP3K1 Thr1402 phosphorylation in neurodegeneration and neuroprotection

  • The antibody has been validated in human brain tissue, making it suitable for neuroscience applications

  • This could reveal new therapeutic targets for neurodegenerative diseases

• Developmental Biology:

  • Mapping the temporal and spatial dynamics of MAP3K1 phosphorylation during embryonic development

  • The antibody's cross-reactivity with human, mouse, and rat samples enables comparative developmental studies

  • This could identify critical signaling events in organogenesis and tissue patterning

• Drug Discovery and Validation:

  • Screening compounds that modulate MAP3K1 Thr1402 phosphorylation

  • The antibody could serve as a pharmacodynamic marker in drug development pipelines

  • This would accelerate development of targeted therapies affecting MAP3K1-dependent pathways

• Single-Cell Signaling Heterogeneity:

  • Investigating cell-to-cell variation in MAP3K1 phosphorylation within tissues

  • The antibody could be adapted for mass cytometry or single-cell Western blotting

  • This would reveal how signaling heterogeneity contributes to tissue function and disease

• Stress Adaptation Mechanisms:

  • Studying how environmental stressors regulate MAP3K1 phosphorylation

  • The antibody could track adaptive versus maladaptive stress responses

  • This would leverage MAP3K1's role in the ERK and JNK kinase pathways that respond to cellular stress

• Immune Cell Signaling Dynamics:

  • Characterizing MAP3K1 phosphorylation in immune cell activation and differentiation

  • The antibody has been validated in Jurkat cells, a model T-cell line

  • This could uncover new regulatory mechanisms in immune response coordination

What reference materials should researchers consult when designing experiments with Phospho-MAP3K1 (Thr1402) Antibody?

Researchers planning experiments with Phospho-MAP3K1 (Thr1402) Antibody should consult these key reference materials:

• Original Research Papers:

  • Schmutz J., et al. (2004) "The DNA sequence and comparative analysis of human chromosome 5," Nature 431:268-274 - Provides genomic context for MAP3K1

  • Xia Y., et al. (1998) "MEK kinase 1 is critically required for c-Jun N-terminal kinase activation by proinflammatory stimuli and growth factor-induced cell migration," Genes Dev. 12:3369-3381 - Establishes functional significance of MAP3K1

  • Vinik B.S., et al. (1995) "MEKK, a major component of the JNK/SAPK activation pathway, is a 196-kDa protein," Mamm. Genome 6:782-783 - Early characterization of MAP3K1

• Technical Resources:

  • Manufacturer's datasheets with application-specific protocols and validation data

  • Antibody validation studies demonstrating specificity in Western blot and immunohistochemistry applications

  • Phosphorylation site databases (e.g., PhosphoSitePlus) for context on Thr1402 conservation and regulation

• Method-Specific Guidelines:

  • For Western blotting: Apply at 1:500-1:1000 dilution

  • For immunohistochemistry: Use at 1:50-1:100 dilution

  • For ELISA applications: Dilute to 1:40000 or 1:1000

• Experimental Controls:

  • Positive control recommendations include Jurkat cells and NIH-3T3 cells

  • Peptide competition controls using the immunizing phospho-peptide sequence around threonine 1402 (T-G-A(p)-G-F)

• Species Considerations:

  • The antibody has validated reactivity with human, mouse, and rat samples

  • Sequence alignment resources to evaluate conservation of the Thr1402 site across species

• Storage and Handling:

  • Store at -20°C or -80°C upon receipt

  • Avoid repeated freeze-thaw cycles that could degrade antibody performance

• Related Signaling Pathway Resources:

  • Literature on the ERK and JNK kinase pathways activated by MAP3K1

  • Information on NF-kappa-B pathway regulation through CHUK and IKBKB activation by MAP3K1