Recombinant Mouse CXCL16 (Extracellular Domain)

Recombinant Mouse CXCL16 (Extracellular Domain) is used in research applications to study its roles in immune regulation, cell signaling, and tissue remodeling, particularly through its interaction with the CXCR6 receptor and its chemokine activity.

CXCL16 exists in both membrane-bound and soluble forms, with the extracellular domain responsible for binding to CXCR6 and mediating chemotactic responses, calcium mobilization, and immune cell recruitment. Using the recombinant extracellular domain allows precise control over experimental conditions, enabling researchers to:

• Investigate immune cell migration and activation: CXCL16 is produced by dendritic cells in lymphoid tissues and acts as a chemoattractant for CXCR6-expressing cells, such as T cells and natural killer T (NKT) cells. Recombinant protein can be used in chemotaxis assays, immune cell activation studies, and cytokine production analyses.
• Study co-stimulatory and antitumor effects: CXCL16 enhances interferon-γ (IFNγ) production in iNKT cells when combined with other stimuli, contributing to improved antitumor responses and immune surveillance. Recombinant CXCL16 can be used to dissect these mechanisms in vitro and in vivo.
• Model tissue remodeling and fibrosis: CXCL16 modulates matrix metalloproteinase activity, collagen synthesis, and fibroblast proliferation, impacting tissue remodeling in heart failure and fibrosis models. Recombinant protein enables mechanistic studies of these processes.
• Elucidate signaling pathways: The extracellular domain of CXCL16 activates intracellular signaling cascades (e.g., JNK, p38 MAPK, ERK) in target cells, which can be studied using recombinant protein in cell-based assays.

Best practices for using recombinant CXCL16 include:

• Employing the extracellular domain to ensure specific interaction with CXCR6 and avoid confounding effects from intracellular or transmembrane regions.
• Using defined concentrations and controls to assess dose-dependent effects on cell migration, activation, or signaling.
• Validating activity through functional assays such as chemotaxis, cytokine release, or receptor binding.

Applications span immunology, oncology, cardiovascular research, and fibrosis studies, making recombinant Mouse CXCL16 (Extracellular Domain) a versatile tool for dissecting chemokine-mediated mechanisms in mouse models and cell systems.

Yes, recombinant Mouse CXCL16 (Extracellular Domain) can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matches the form of CXCL16 detected by your assay.

Essential context and supporting details:

• ELISA standards are commonly recombinant proteins, including CXCL16, and are used to generate standard curves for quantification. The standard should be of the same species and domain as the analyte measured in your samples.
• The extracellular domain is typically the relevant portion for soluble CXCL16 detected in most sandwich ELISA formats, which measure the secreted or shed form in serum, plasma, or cell culture supernatants.
• Validation is critical: Ensure the recombinant protein is pure, correctly folded, and its concentration is accurately determined. The standard curve should be linear and reproducible within the assay’s dynamic range.
• Expression system and formulation: Recombinant standards may be produced in E. coli, yeast, or mammalian cells. The expression system can affect post-translational modifications, which may influence antibody recognition. If your ELISA kit uses antibodies that recognize the native glycosylation or other modifications, confirm that your recombinant standard is compatible.
• Carrier proteins (e.g., BSA): Some recombinant proteins are supplied with carrier proteins to stabilize them. If your standard is carrier-free, ensure it is handled carefully to avoid adsorption losses.
• Preparation: Reconstitute and dilute the recombinant standard according to best practices, using the same buffer as recommended for the ELISA kit to minimize matrix effects.

Best practices:

• Run a full standard curve with your recombinant CXCL16 in parallel with your samples for each assay.
• Confirm that the standard curve is linear and covers the expected concentration range of your samples.
• If switching from a kit-provided standard to your own recombinant protein, validate equivalency by comparing standard curves and recovery in spiked samples.

Limitations:

• If your recombinant CXCL16 differs in sequence, post-translational modification, or folding from the native protein in your samples, quantification may be affected. Always verify compatibility with your specific ELISA antibodies and detection system.

In summary, recombinant Mouse CXCL16 (Extracellular Domain) is suitable as an ELISA standard if it is validated for your assay format and matches the analyte detected by your antibodies.

Recombinant Mouse CXCL16 (Extracellular Domain) has been validated for several applications in published research, primarily in the context of immunology, chemotaxis, and functional assays.

Key validated applications include:

• Chemotaxis/Cell Migration Assays: The extracellular domain of CXCL16 functions as a chemoattractant for cells expressing the CXCR6 receptor, such as activated T cells and certain lymphocyte subsets. Published studies have used recombinant mouse CXCL16 to demonstrate its ability to induce chemotactic responses in vitro and in vivo, particularly in immune cell recruitment models.

• Functional Bioactivity Assays: Recombinant mouse CXCL16 has been validated in bioactivity assays to confirm its ability to activate CXCR6+ cells, induce calcium mobilization, and modulate immune responses. These assays often measure downstream signaling events or cellular responses following CXCL16 stimulation.

• ELISA (Enzyme-Linked Immunosorbent Assay): The protein has been validated as a standard or control in ELISA-based quantification of CXCL16 or for detection of anti-CXCL16 antibodies.

• Receptor Binding Studies: The extracellular domain is used to study binding interactions with its receptor CXCR6, as well as its function as a scavenger receptor for oxidized low-density lipoprotein (OxLDL) on macrophages.

• In Vivo Models: Recombinant CXCL16 has been used in mouse models to study its role in immune cell recruitment, tumor immunity, and atherosclerosis, often by administering the protein or expressing it in engineered systems to assess physiological effects.

• Inflammation and Disease Models: Research has validated its use in models of inflammation, tissue injury, fibrosis, and neuroinflammation, where CXCL16/CXCR6 axis involvement is investigated.

Summary Table of Validated Applications

Additional Notes:

• The extracellular domain is the active, soluble form responsible for chemokine activity, distinct from the full-length transmembrane protein.
• Published research often uses recombinant CXCL16 to dissect its dual roles as a chemokine and scavenger receptor, particularly in macrophage biology and cardiovascular disease.

If you require protocols or specific assay details for any of these applications, please specify the intended use.

To reconstitute and prepare Recombinant Mouse CXCL16 (Extracellular Domain) protein for cell culture experiments, dissolve the lyophilized protein at 100 μg/mL in sterile PBS under aseptic conditions.

Essential steps and details:

• Reconstitution:

  • Add sterile PBS directly to the lyophilized protein to achieve a final concentration of 100 μg/mL.
  • Gently mix by pipetting or swirling; avoid vigorous agitation to prevent protein denaturation.
  • If recommended by the supplier, you may also use sterile ultrapure water (≥18 MΩ·cm) for initial dissolution, but PBS is preferred for physiological compatibility.

• Aliquoting and Storage:

  • After reconstitution, aliquot the solution into small volumes to avoid repeated freeze-thaw cycles, which can degrade the protein.
  • Store aliquots at −20 °C to −70 °C in a manual defrost freezer for up to 3 months.
  • For short-term use (up to 1 month), storage at 2–8 °C is acceptable.

• Preparation for Cell Culture:

  • Before use, dilute the stock solution to the desired working concentration using sterile cell culture medium or PBS.
  • Typical working concentrations for chemotaxis or signaling assays range from 4–20 ng/mL, based on reported ED50 values for CXCL16 activity.
  • Ensure the final buffer is compatible with your cell culture system and does not contain additives that may affect cell viability or protein function.

• Handling Precautions:

  • Maintain sterility throughout preparation to prevent contamination.
  • Avoid repeated freeze-thaw cycles by using aliquots.
  • If required, supplement the dilution buffer with carrier proteins (e.g., 0.1% BSA) to minimize adsorption to plasticware, especially at low concentrations.

Summary Table: Recombinant Mouse CXCL16 Preparation

Additional notes:

• Confirm the endotoxin level is suitable for cell culture (<0.1 EU/μg is typical for recombinant proteins).
• Always consult the specific Certificate of Analysis for your batch for any additional instructions.

This protocol ensures optimal solubility, stability, and biological activity of recombinant CXCL16 for cell-based assays.