Recombinant Rat MCP-1

Recombinant Rat MCP-1 is widely used in research applications to study immune cell recruitment, inflammation, and disease mechanisms due to its well-characterized role as a chemokine that specifically attracts monocytes and other immune cells. Using the recombinant form ensures experimental consistency, purity, and the ability to control for variables such as endotoxin contamination.

Key scientific applications and advantages include:

• Cell Migration and Chemotaxis Assays: MCP-1 is a potent chemoattractant for monocytes, basophils, and dendritic cells, making recombinant rat MCP-1 essential for in vitro and in vivo studies of cell migration, immune cell recruitment, and chemotactic signaling pathways.

• Inflammation and Disease Models: MCP-1 is upregulated in response to tissue injury and inflammation. Recombinant rat MCP-1 is used to model inflammatory responses, study mechanisms of vascular injury, and investigate the development of diseases such as atherosclerosis, neuropathic pain, and CNS injury.

• Blocking and Specificity Controls: Recombinant MCP-1 serves as a ligand-blocking control in immunofluorescent staining and flow cytometry, allowing researchers to confirm the specificity of MCP-1 detection and antibody binding.

• Mechanistic Studies: MCP-1 influences vascular smooth muscle cell (VSMC) behavior, neuronal activation, and microglial chemotaxis. Recombinant protein enables precise mechanistic studies of MCP-1/CCR2 signaling, cell polarization, and downstream effects such as Rac1 activation.

• Therapeutic Research: MCP-1 is a target for therapeutic intervention in inflammatory and autoimmune diseases. Recombinant rat MCP-1 is used to test neutralizing antibodies, antisense oligonucleotides, and other inhibitors in preclinical models.

• Standardization and Reproducibility: Recombinant proteins provide defined concentration, purity, and activity, reducing variability compared to native or crude preparations. This is critical for quantitative assays and reproducible results.

Best practices when using recombinant rat MCP-1:

• Confirm protein purity and low endotoxin levels to avoid confounding immune activation.
• Use appropriate controls (e.g., blocking controls, neutralizing antibodies) to validate specificity in functional assays.
• Optimize concentration and exposure time for your specific cell type or animal model, as MCP-1 effects are dose- and time-dependent.

In summary, recombinant rat MCP-1 is a versatile tool for dissecting immune cell behavior, inflammatory mechanisms, and therapeutic interventions in rat models, with broad utility in immunology, neuroscience, and vascular biology research.

Yes, recombinant rat MCP-1 can be used as a standard for quantification or calibration in ELISA assays, provided it is validated for this purpose in your specific assay system. Recombinant MCP-1 is commonly used as a quantitative standard in sandwich ELISA protocols for measuring rat MCP-1 protein levels.

Key considerations and supporting details:

• Intended Use: Recombinant rat MCP-1 is routinely supplied as a standard in commercial ELISA kits and is specifically recommended for generating standard curves to quantify MCP-1 in biological samples.
• Assay Compatibility: Most rat MCP-1 ELISA kits are designed to recognize both natural and recombinant forms of MCP-1, and standard curves generated with recombinant protein are used to interpolate sample concentrations.
• Validation: It is important to ensure that the recombinant MCP-1 you use is of high purity and properly reconstituted according to the manufacturer’s instructions. The standard should be validated for parallelism with endogenous MCP-1 in your sample matrix to ensure accurate quantification.
• Calibration: The standard curve should be prepared by serial dilution of the recombinant MCP-1 in the same buffer or diluent used for your samples, as specified in your ELISA protocol.
• Research Use: Most recombinant MCP-1 standards and ELISA kits are for research use only and not for diagnostic applications.

Best Practices:

• Always check that the recombinant standard matches the sequence and post-translational modifications (if relevant) of the endogenous protein in your samples.
• Confirm that your ELISA kit or custom assay has been validated to detect the recombinant form equivalently to the native protein.
• Prepare and store the standard according to the datasheet to maintain stability and activity.

Summary:
You can use recombinant rat MCP-1 as a standard for ELISA quantification, as long as it is validated for your assay and you follow best practices for preparation and calibration.

Recombinant Rat MCP-1 (CCL2) has been validated in published research for a range of applications, primarily involving immune cell recruitment, chemotaxis, pain models, and as a standard or control in immunoassays.

Key validated applications include:

• Bioactivity assays: Used to demonstrate chemotactic activity for monocytes, T cells, and dendritic cells in vitro and in vivo, as well as to stimulate migration of fibroblasts and other cell types.
• ELISA standard: Employed as a quantitative standard for measuring MCP-1 levels in biological samples using ELISA kits.
• Western blot control: Used as a positive control for MCP-1 detection in Western blotting.
• Neutralization assays: Utilized to assess the efficacy of neutralizing antibodies or inhibitors targeting MCP-1.
• In vivo functional studies: Injected into animal models to study its role in monocyte/macrophage recruitment, inflammation, pain hypersensitivity, and tissue injury responses.
• Cellular bioassays: Applied to cultured cells (e.g., microglia, fibroblasts, sensory neurons) to assess downstream signaling, activation, and migration.
• Flow cytometry (intracellular block): Used in flow cytometry protocols to block or detect MCP-1 in cells.

Representative published research applications:

• Pain and neuroinflammation: Recombinant rat MCP-1 has been injected into muscle or nervous tissue to induce and study mechanical hyperalgesia and neuronal activation, supporting its role in pain pathways.
• Immune cell migration: Used in transwell and in vivo migration assays to demonstrate chemotactic effects on monocytes, T cells, and macrophages.
• Cardiovascular disease models: Applied in studies of heart failure to assess MCP-1 expression and binding in rat myocardium, implicating it in monocyte/macrophage infiltration and cardiac remodeling.
• Inflammation and tissue injury: Used to model and analyze inflammatory responses, including macrophage recruitment in testicular inflammation and wound healing.
• Tumor biology: Studied for its role in tumor progression and angiogenesis by promoting immune cell infiltration and neovascularization.

Summary Table:

These applications are well-supported by both product validation data and peer-reviewed research, demonstrating the versatility of recombinant rat MCP-1 in immunology, neuroscience, cardiovascular, and cancer biology studies.

To reconstitute and prepare Recombinant Rat MCP-1 protein for cell culture experiments, briefly centrifuge the vial to collect the lyophilized powder, then reconstitute in sterile water or PBS at a concentration of 0.1–1.0 mg/mL, optionally including 0.1–1% carrier protein (such as BSA) to prevent adsorption and loss of activity.

Essential steps and considerations:

• Centrifuge the vial briefly before opening to ensure all powder is at the bottom.
• Reconstitution buffer: Use sterile distilled water or PBS. For cell culture, PBS is preferred, and adding 0.1–1% BSA (bovine serum albumin) is recommended to stabilize the protein and minimize loss due to adsorption to plasticware, especially if working in serum-free conditions.
• Concentration: Typical reconstitution concentrations range from 0.1 mg/mL to 1.0 mg/mL. For most applications, 100 μg/mL is commonly used.
• Mixing: Gently pipette the solution to dissolve the protein. Do not vortex, as vigorous mixing can denature the protein.
• Aliquoting: After reconstitution, aliquot the solution to avoid repeated freeze-thaw cycles, which can degrade the protein.
• Storage: Store aliquots at −20°C or −80°C for long-term use. Short-term storage (up to one week) can be at 4°C.
• Dilution for cell culture: Dilute the reconstituted stock into your cell culture medium to the desired working concentration. If using serum-free medium, ensure the presence of carrier protein (e.g., 0.1–1% BSA) in all solutions to prevent loss of MCP-1.

Protocol summary:

1. Briefly centrifuge the vial.
2. Add sterile PBS (with 0.1–1% BSA) to achieve 0.1–1.0 mg/mL.
3. Gently pipette to dissolve; do not vortex.
4. Aliquot and store at −20°C or −80°C.
5. Dilute into cell culture medium as needed.

Additional notes:

• Avoid repeated freeze-thaw cycles.
• Always use sterile technique to prevent contamination.
• For bioassays, confirm the final concentration and activity using appropriate controls.

This protocol ensures optimal recovery, stability, and biological activity of recombinant rat MCP-1 for cell culture experiments.