Recombinant Mouse MCP-2

Recombinant Mouse MCP-2 (CCL8) is used in research applications to study immune cell recruitment, inflammatory responses, and chemokine signaling due to its role as a potent chemoattractant for monocytes, lymphocytes, eosinophils, and basophils.

Key scientific reasons to use recombinant Mouse MCP-2 in your research include:

• Modeling Inflammatory and Allergic Responses: MCP-2 is a cytokine critical for mediating allergic and inflammatory processes. It activates mast cells, eosinophils, and basophils, and is involved in recruiting monocytes to sites of inflammation, making it valuable for studying mechanisms of immune cell trafficking and inflammation in mouse models.

• Chemotaxis Assays: Recombinant MCP-2 is commonly used to induce chemotaxis in vitro, particularly for monocytes and THP-1 cells, allowing quantification of cell migration in response to chemokine gradients. This is essential for dissecting chemokine-receptor interactions and leukocyte behavior.

• Receptor Specificity Studies: MCP-2 uniquely signals through multiple chemokine receptors (CCR1, CCR2B, and CCR5), unlike other related chemokines. This property enables detailed studies of receptor-ligand specificity and downstream signaling pathways.

• Comparative Immunology and Preclinical Evaluation: Recombinant MCP-2 is used as a standardized reagent in mouse model studies, comparative immunology, and preclinical therapeutic evaluations, facilitating reproducible and controlled experiments.

• Tumor Microenvironment and Antiviral Research: MCP-2 contributes to tumor-associated leukocyte infiltration and has been implicated in the antiviral state, including as a natural inhibitor of HIV-1 via CCR5 signaling, making it relevant for cancer and infectious disease research.

• Quality and Consistency: Recombinant forms are typically highly purified, endotoxin-controlled, and biologically active, ensuring reproducibility and minimizing variability compared to native or crude preparations.

Typical applications include:

• In vitro chemotaxis and migration assays
• In vivo models of inflammation, allergy, or tumor immunology
• Receptor binding and signaling studies
• Functional assays for immune cell activation and proliferation

In summary, using recombinant Mouse MCP-2 provides a reliable, well-characterized tool for dissecting chemokine biology, immune cell dynamics, and inflammatory mechanisms in mouse-based research systems.

Recombinant Mouse MCP-2 (CCL8) can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to the assay system. This is a common practice in cytokine and chemokine quantification, as recombinant proteins offer defined concentrations and high purity for standard curve generation.

Key considerations and best practices:

• Assay Compatibility: The recombinant MCP-2 must be recognized by the antibodies used in your ELISA kit. Most commercial Mouse MCP-2 ELISA kits are designed to detect both natural and recombinant MCP-2, and their protocols typically include recombinant MCP-2 as the standard for calibration.
• Standard Preparation: Prepare a serial dilution of the recombinant MCP-2 in the same buffer or diluent recommended by your ELISA kit protocol to generate a standard curve. This allows interpolation of sample concentrations based on the standard curve.
• Validation: It is important to validate that the recombinant MCP-2 behaves similarly to endogenous MCP-2 in your sample matrix. Recovery experiments (spiking known amounts into sample matrices) are recommended to confirm accuracy and parallelism of the standard curve.
• Intended Use: Ensure the recombinant MCP-2 is for research use only and not for diagnostic procedures, as specified by most kit manufacturers.
• Bioactivity: While recombinant MCP-2 is suitable for ELISA calibration, it may not be validated for bioassays unless specifically tested for biological activity.

Protocol Example:

1. Reconstitute recombinant MCP-2 according to the supplier’s instructions.
2. Prepare serial dilutions (e.g., 1000, 500, 250, 125, 62.5, 31.25, 15.6 pg/mL) in the recommended diluent.
3. Run the ELISA according to kit instructions, using the recombinant MCP-2 dilutions to generate the standard curve.
4. Quantify unknown samples by interpolating their absorbance values against the standard curve.

Limitations:

• Do not mix standards or reagents from different kits or vendors, as performance cannot be guaranteed.
• Recombinant standards should be freshly prepared and used promptly to ensure stability and accuracy.

Summary Table:

In conclusion, recombinant Mouse MCP-2 is widely accepted as a standard for ELISA quantification, provided it is compatible with your assay and validated for accuracy in your sample matrix.

Recombinant Mouse MCP-2 (CCL8) has been validated for several key applications in published research, primarily in the context of immunology and cell biology.

The main validated applications include:

• ELISA (Enzyme-Linked Immunosorbent Assay): Used to quantify MCP-2 levels in biological samples, supporting studies of cytokine secretion and immune responses.
• Functional Assays (e.g., Chemotaxis): MCP-2 is widely used to assess its ability to induce chemotactic migration of immune cells such as monocytes, lymphocytes, basophils, eosinophils, and Th2 cells. These assays demonstrate the protein’s biological activity and its role in recruiting leukocytes to sites of inflammation.
• Western Blot: Used to detect MCP-2 protein in cell lysates or tissue extracts, confirming expression and molecular weight.
• Immunohistochemistry: Applied to tissue sections to localize MCP-2 expression in situ, often in studies of inflammation or immune cell infiltration.

Supporting details:

• MCP-2/CCL8 is a chemokine involved in the recruitment of various leukocyte populations during inflammatory and allergic responses.
• Functional validation often involves chemotaxis assays, where recombinant MCP-2 is shown to induce migration of specific immune cell subsets, such as Th2 cells via the CCR8 receptor.
• ELISA and Western blot validations confirm the protein’s detectability and specificity in complex biological samples.
• Immunohistochemistry enables spatial mapping of MCP-2 expression in tissues, supporting studies of its role in disease models.

Additional notes:

• These applications are for research use only and not for clinical or diagnostic purposes.
• Published research frequently uses recombinant mouse MCP-2 to dissect mechanisms of immune cell trafficking and to model inflammatory diseases in vitro and in vivo.

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

To reconstitute and prepare Recombinant Mouse MCP-2 protein for cell culture experiments, briefly centrifuge the vial to collect the lyophilized powder at the bottom, then add sterile, distilled water to achieve a concentration of 0.1–1.0 mg/mL. Gently pipette to dissolve the protein, avoiding vigorous mixing or vortexing.

Detailed protocol:

• Centrifugation: Spin the vial briefly before opening to ensure all powder is at the bottom.
• Reconstitution: Add sterile, distilled water (pH 7–8 is typical; some protocols specify pH 8.0) to reach a final concentration of 0.1–1.0 mg/mL. For example, add 100 µL water to 100 µg protein for 1 mg/mL, or 1 mL for 0.1 mg/mL.
• Dissolution: Gently pipette up and down, washing the sides of the vial to ensure complete dissolution. Do not vortex.
• Carrier protein: For further dilution, use an aqueous buffer containing 0.1–1.0% BSA (bovine serum albumin) or another carrier protein to stabilize MCP-2 and minimize adsorption to plasticware.
• Aliquoting and storage: After reconstitution, aliquot the solution to avoid repeated freeze-thaw cycles. Store aliquots at -80°C for long-term storage, or at 4°C for short-term use (up to one week). For long-term storage, always include carrier protein in the buffer.
• Working concentration: For cell culture experiments, MCP-2 is typically used at 1–100 ng/mL, depending on the assay and cell type. Prepare working solutions by diluting the stock in cell culture medium or buffer containing carrier protein.

Best practices:

• Use sterile technique throughout to prevent contamination.
• Avoid repeated freeze-thaw cycles by aliquoting.
• If the protein is sensitive, minimize exposure to room temperature and light.
• Confirm biological activity with a chemotaxis assay or other functional readout if required.

Summary Table: Recombinant Mouse MCP-2 Reconstitution

This protocol ensures optimal solubility, stability, and biological activity of recombinant Mouse MCP-2 for cell culture applications.