Recombinant Mouse Axl

Recombinant mouse AXL is a valuable tool for research applications across multiple biological domains, offering both mechanistic insights and functional validation opportunities.

Key Research Applications

Immune Cell Signaling Studies

Recombinant mouse AXL enables investigation of receptor tyrosine kinase signaling in immune cells, where AXL is expressed on dendritic cells, macrophages, and regulatory T cells. This is particularly useful for studying how AXL modulates immune responses, including its role in dampening inflammatory reactions, regulating cytokine secretion, and facilitating the clearance of apoptotic cells and debris.

Gas6/AXL Axis Investigation

Using recombinant AXL allows you to examine the Gas6/AXL signaling pathway, which plays critical roles in multiple physiological contexts. Studies have demonstrated that recombinant Gas6 restores phosphorylated AXL expression in a dose-dependent manner, making recombinant AXL proteins essential for characterizing ligand-receptor interactions and downstream signaling cascades.

Disease Model Research

Recombinant mouse AXL is instrumental for studying AXL's involvement in various pathological conditions. The protein is upregulated in disease states such as demyelination models and multiple sclerosis lesions, making it valuable for investigating AXL's role in neuroinflammation and disease progression. Additionally, AXL's involvement in cardiac repair following ischemic injury and alveolar inflammation during ischemia-reperfusion injury makes recombinant AXL useful for cardiovascular and pulmonary research.

Cancer Biology and Metastasis Studies

AXL expression correlates with poor prognosis in several cancers and mediates oncogenic phenotypes including cell survival, migration, invasion, and metastasis. Recombinant AXL enables functional studies examining how AXL activation promotes chemoresistance and contributes to epithelial-mesenchymal transition (EMT) processes.

Therapeutic Target Validation

Recombinant mouse AXL serves as a critical tool for validating AXL as a therapeutic target. By using recombinant protein in functional assays, you can assess how AXL inhibition affects immune cell function, antiviral responses, and tumor immunity, supporting the development and optimization of AXL-targeted therapeutics.

The availability of recombinant mouse AXL with various tags and fusion formats (such as His-tagged or Fc-chimera constructs) provides flexibility for different experimental approaches, including binding assays, cell stimulation experiments, and structural studies.

Yes, you can use recombinant mouse Axl as a standard for quantification and calibration in ELISA assays, but there are important considerations and limitations to understand.

Suitability for ELISA Standards

Recombinant mouse Axl is appropriate for use as an ELISA standard. The assay format is specifically designed to recognize both natural and recombinant forms of the protein, making recombinant versions suitable for standard curve preparation. Many commercial ELISA kits for mouse Axl include recombinant protein standards as part of their reagent packages.

Key Considerations for Accurate Calibration

Protein Purity and Characterization

When preparing standard curves, a purified protein should ideally be used. If using recombinant protein that requires semi-purification in your laboratory, you should measure the concentration using high-performance liquid chromatography (HPLC) or another validated quantitation method to ensure accurate concentration determination.

Carrier Selection

Recombinant proteins are available in different formulations. For use as ELISA standards, recombinant mouse Axl with carrier proteins (such as BSA) is recommended. Carrier-free formulations are typically not recommended for ELISA calibration applications, as they may behave differently in the assay system.

Dilution Accuracy

A critical challenge when using recombinant proteins as QC controls or standards involves large dilution steps. Typical dilutions range from micrograms per milliliter to picograms per milliliter. These substantial dilutions can introduce cumulative errors, and the larger the dilution factor, the greater the potential for inaccuracy in your final measurements.

Standard Curve Validation

Do not extend the standard curve beyond the validated range. The standard curve range has been validated for reliable performance across multiple kit lots and operators. Adding concentrations above or below the established range may result in inconsistent signals or false positives.

Best Practices

For optimal results, run internal controls alongside your standard curve and blanks. These controls should duplicate the composition of your actual samples as closely as possible, allowing you to verify that the same analyte concentration produces consistent results in both the standard diluent and your sample matrix.

Recombinant Mouse Axl has been validated for several key applications in published research, primarily in immunization for antibody generation, ELISA, and functional assays related to cancer biology and receptor-ligand interactions.

Key validated applications include:

• Immunogen for Antibody Generation: Recombinant mouse Axl has been used as an immunogen to generate Axl-specific single domain antibodies (sdAbs) in alpacas. These sdAbs were then used for diagnostic and therapeutic studies in acute myeloid leukemia (AML). The recombinant protein was critical for both immunization and subsequent screening of antibody clones.

• ELISA (Enzyme-Linked Immunosorbent Assay): The recombinant protein has been used as a coating antigen in ELISA to screen for antibody binding and to confirm the specificity and functionality of generated antibodies. Radio-ELISA was also performed using recombinant mouse Axl to quantify antibody-antigen interactions.

• Functional Assays: Recombinant mouse Axl has been used in in vitro assays to assess the biological activity of antibodies, including viability, apoptosis, and proliferation assays in cancer cell lines. These assays help determine the therapeutic potential of Axl-targeted agents.

• Phage Display and Biopanning: The protein has been used as a target in phage display and biopanning protocols to select high-affinity antibody clones.

• Receptor-Ligand Interaction Studies: Recombinant Axl is commonly used to study its interaction with ligands such as Gas6, which is relevant for understanding signaling pathways in cancer and immune regulation.

• Standard/Control in Analytical Assays: Recombinant mouse Axl is used as a standard or positive control in various analytical assays, including Western blot and ELISA, to validate antibody specificity and assay performance.

Additional research contexts:

• Cancer Biology: Recombinant Axl is widely used to investigate its role in tumor progression, drug resistance, and as a therapeutic target in various cancers.
• Viral Entry Studies: While most published work focuses on human Axl, recombinant Axl proteins (including mouse) are used to study viral receptor functions, such as in SARS-CoV-2 research.

Summary Table: Applications of Recombinant Mouse Axl in Published Research

Note: The specific recombinant mouse Axl protein used in these studies is typically expressed in mammalian cells (e.g., HEK293), ensuring proper folding and post-translational modifications relevant for functional studies.

If you need protocols or more technical details for a specific application, please specify the intended use.

To reconstitute and prepare Recombinant Mouse Axl protein for cell culture experiments, dissolve the lyophilized protein in sterile distilled water or PBS, aiming for a concentration between 0.1–1.0 mg/mL, depending on your experimental requirements.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to ensure all lyophilized material is at the bottom.
• Add sterile distilled water or PBS (pH 7.4) directly to the vial. For most applications, a concentration of 0.5 mg/mL is recommended if the total amount is 100 μg or less. For larger amounts, adjust the volume accordingly.
• Gently mix by swirling or inverting the vial. Avoid vigorous pipetting or vortexing, as this can denature the protein.
• Allow the protein to fully dissolve at room temperature for 10–30 minutes. If necessary, gently pipette up and down to aid dissolution, but do not create bubbles.
• Aliquot the reconstituted protein into single-use volumes to avoid repeated freeze-thaw cycles, which can degrade the protein.
• Storage after reconstitution:
  • Store aliquots at –80°C for long-term use (up to 3 months).
  • For short-term use (within 1 week), store at 2–8°C.
  • Avoid repeated freeze-thaw cycles.

Additional considerations:

• The lyophilized protein is typically formulated with trehalose or mannitol as a stabilizer.
• If your application is sensitive to buffer components, dialyze or exchange the buffer after reconstitution.
• For cell culture, ensure the final working solution is sterile. If necessary, filter the reconstituted protein through a 0.22 μm filter before use.
• If increased stability is needed, adding 5–50% glycerol to aliquots before freezing can help.

Summary Table:

Always consult the specific product’s Certificate of Analysis or data sheet for any lot-specific instructions.