Recombinant Human MCP-4

Recombinant Human MCP-4 (CCL13) is used in research applications to study immune cell recruitment, inflammation, and allergic responses due to its role as a potent chemoattractant for monocytes, eosinophils, basophils, and T lymphocytes.

MCP-4 is a member of the CC chemokine family and is induced by inflammatory cytokines such as IL-1 and TNF-α. Its primary scientific applications include:

• Chemotaxis Assays: MCP-4 is widely used to induce and measure the migration of immune cells (e.g., monocytes, eosinophils, T cells) in vitro, which is critical for dissecting mechanisms of immune cell trafficking in inflammation and allergy models.
• Receptor Studies: MCP-4 binds to and activates several chemokine receptors (CCR2, CCR3, and CCR5), making it valuable for receptor-ligand interaction studies and for screening receptor agonists or antagonists.
• Disease Modeling: Elevated MCP-4 expression is implicated in various inflammatory diseases, such as rheumatoid arthritis and renal inflammation, making recombinant MCP-4 useful for modeling these conditions and evaluating potential therapeutic interventions.
• Cell Signaling Research: MCP-4 can be used to stimulate signaling pathways in target cells, enabling the study of downstream effects such as calcium flux, cytokine release, and cell proliferation.
• Allergy and Asthma Research: MCP-4’s ability to recruit eosinophils and basophils is particularly relevant for studying allergic inflammation and asthma pathogenesis.

Key advantages of using recombinant MCP-4:

• Defined, consistent activity: Recombinant proteins provide batch-to-batch consistency and eliminate variability associated with native protein isolation.
• Species specificity: Human MCP-4 is required for studies involving human cells or receptors to ensure physiological relevance.
• Versatility: Suitable for a range of applications, including chemotaxis, receptor binding, and cell signaling assays.

In summary, recombinant human MCP-4 is a critical tool for investigating immune cell migration, inflammatory mechanisms, and chemokine receptor biology in human disease models.

Yes, recombinant human MCP-4 can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and its concentration is accurately known. This is a common and accepted practice in quantitative ELISA protocols.

Supporting context and best practices:

• Recombinant proteins are widely used as ELISA standards: Most commercial MCP-4 ELISA kits use recombinant human MCP-4 as the standard to generate the calibration curve for quantification of MCP-4 in biological samples. The standard curve is essential for interpolating the concentration of MCP-4 in unknown samples.

• Purity and quantification: For best results, the recombinant MCP-4 should be highly purified and its concentration should be determined accurately, typically by absorbance at 280 nm or another validated method. Impurities or inaccurate quantification can introduce error into your standard curve.

• Matrix effects: If your samples are in a complex biological matrix (e.g., serum or plasma), ensure that the recombinant standard is prepared in a similar matrix or with appropriate diluents to minimize matrix effects and improve accuracy.

• Validation: It is important to validate that your recombinant MCP-4 behaves similarly to native MCP-4 in your assay system. Some kits or protocols may note that recombinant proteins can behave differently due to post-translational modifications or folding differences, but for most sandwich ELISAs, recombinant standards are the norm and are validated for this use.

• Protocol adherence: Follow the reconstitution and dilution instructions provided with your recombinant MCP-4 or ELISA kit, as these can be lot-specific and affect the accuracy of your standard curve.

Summary of key points:

• Recombinant human MCP-4 is suitable and commonly used as an ELISA standard for quantification and calibration.
• Ensure high purity and accurate quantification of the recombinant protein.
• Prepare standards in a matrix similar to your samples to minimize matrix effects.
• Validate equivalence to native protein if possible, especially if using custom or in-house recombinant preparations.

If you are using a commercial ELISA kit, the included recombinant MCP-4 standard is specifically validated for this purpose and should be used as directed by the kit protocol. If you are developing your own assay, ensure your recombinant standard meets the above criteria for reliable quantification.

Recombinant Human MCP-4 (CCL13) has been validated in published research for applications including bioassays (functional cell-based assays), ELISA (as a standard), and chemotaxis assays.

Key validated applications in the literature:

• Bioassays: MCP-4 is widely used to assess its ability to chemoattract various cell types, including monocytes, eosinophils, basophils, and T lymphocytes. These assays typically measure cell migration or activation in response to MCP-4, confirming its functional activity through receptor-mediated signaling.

• ELISA (Enzyme-Linked Immunosorbent Assay): Recombinant MCP-4 is used as a standard in ELISA protocols to quantify MCP-4 levels in biological samples, such as cell culture supernatants or patient specimens. This application is essential for studies investigating MCP-4 expression in disease states or immune responses.

• Chemotaxis Assays: MCP-4’s primary biological activity is as a chemoattractant. Published studies have validated its use in chemotaxis assays to measure directed migration of monocytes and other leukocytes.

• Cell Signaling Studies: MCP-4 has been used to activate signaling pathways in monocytes, T lymphocytes, eosinophils, and basophils, particularly in the context of inflammation and allergic responses. These studies often involve measuring downstream effects such as cytokine release or receptor activation.

• Cancer Research: MCP-4 has been implicated in cancer progression, notably ovarian cancer, where it was shown to trigger epithelial-mesenchymal transition and promote tumor growth via the p38 MAPK pathway. Functional assays in these studies validate MCP-4’s role in cell signaling and migration.

Summary Table of Validated Applications

Additional Notes:

• MCP-4 is typically validated using whole cells or transfected cell lines expressing relevant chemokine receptors (CCR2, CCR3, CCR5).
• Its use as an ELISA standard is well-established for quantifying MCP-4 in immunological and clinical research.
• Functional validation often includes dose-response studies to determine effective concentrations for chemotaxis or activation.

If you require protocol details or specific assay conditions, please specify the application of interest.

To reconstitute and prepare Recombinant Human MCP-4 (CCL13) protein for cell culture experiments, first centrifuge the vial briefly to collect the lyophilized powder at the bottom. Then, add sterile distilled water or sterile PBS (pH 7.2–7.4) to achieve a concentration between 0.1–1.0 mg/mL; a commonly used starting point is 0.1 mg/mL. Gently pipette the solution down the sides of the vial to dissolve the protein, avoiding vigorous agitation or foaming.

Key steps and best practices:

• Centrifuge the vial before opening to ensure all powder is at the bottom.
• Reconstitute in sterile distilled water or PBS (pH 7.2–7.4) to a concentration of 0.1–1.0 mg/mL.
• For enhanced stability and to minimize adsorption, you may add 0.1% low endotoxin BSA or heat-inactivated fetal calf serum to the buffer for further dilutions.
• Allow the protein to dissolve at room temperature for 15–30 minutes with gentle agitation.
• Avoid repeated freeze-thaw cycles; aliquot the reconstituted protein and store at −20°C to −70°C for long-term storage, or at 2–8°C for up to one month.
• For cell culture, dilute the stock solution to the desired working concentration using tissue culture-grade buffer or medium containing carrier protein (e.g., BSA or serum).

Summary Table: Reconstitution and Handling

Always consult the specific product datasheet for any unique formulation or handling requirements, as minor differences may exist between preparations.