Recombinant Mouse M-CSF

Recombinant mouse M-CSF (macrophage colony-stimulating factor) is a valuable tool for research applications due to its critical role in myeloid cell biology and its well-characterized functional properties.

Key Biological Functions

Recombinant mouse M-CSF serves as a potent growth factor and differentiation agent for myeloid lineage cells. It stimulates the proliferation and differentiation of hematopoietic stem cells into monocyte and macrophage populations in culture, making it essential for generating these cell types in vitro. The protein regulates the differentiation, proliferation, and survival of osteoclasts, macrophages, monocytes, and their hematopoietic progenitors.

Enhanced Cellular Functions

Beyond differentiation, recombinant mouse M-CSF enhances multiple functional capabilities of myeloid cells. It promotes the survival and activation of macrophages while enhancing their phagocytic activity, cytokine production, and antigen presentation capabilities. The protein also increases cytotoxicity, superoxide production, phagocytosis, chemotaxis, and secondary cytokine production in monocytes and macrophages. Additionally, M-CSF primes macrophages for improved killing of tumor cells and microorganisms, and stimulates pinocytosis.

Research Applications

Recombinant mouse M-CSF has been optimized for use in cell culture, differentiation studies, and functional assays. It binds to the colony stimulating factor 1 receptor (c-fms) and stimulates cell proliferation in established cell lines in a dose-dependent manner. The protein is particularly valuable for studies involving macrophage biology, immune responses, bone metabolism, and inflammatory processes.

Therapeutic Research Potential

The protein's role in immune regulation makes it relevant for investigating therapeutic strategies. M-CSF modulation has shown promise in research contexts for inflammatory and autoimmune diseases, as well as certain cancers. Additionally, studies have demonstrated that M-CSF treatment can enhance protection against bacterial and fungal infections by promoting myeloid cell development and function, making it useful for immunological research applications.

Yes, recombinant Mouse M-CSF can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and bioactivity, and is compatible with the antibodies and detection system used in your assay.

Supporting details:

• Intended Use: Multiple sources explicitly state that recombinant mouse M-CSF is suitable as an ELISA standard. For example, one supplier notes their recombinant M-CSF is "ideal for cell culture, ELISA standards, and Western Blot controls". ELISA kits from several manufacturers use recombinant mouse M-CSF as the standard for generating calibration curves.
• Assay Compatibility: Commercial ELISA kits for mouse M-CSF are validated to recognize both natural and recombinant forms of the protein, and their standard curves are generated using recombinant M-CSF. The Quantikine ELISA, for example, is calibrated against highly purified recombinant mouse M-CSF and demonstrates parallelism between natural and recombinant forms.
• Expression System Considerations: The recombinant M-CSF should closely mimic the native protein in terms of folding and post-translational modifications. Proteins expressed in eukaryotic systems (e.g., yeast, HEK293) are generally preferred for standards, as they more closely resemble the native protein compared to those expressed in E. coli.
• Reconstitution and Handling: Follow the manufacturer’s instructions for reconstitution (often in PBS with carrier protein) and ensure the standard is prepared fresh for each assay to maintain accuracy.
• Validation: If using a recombinant M-CSF preparation not supplied with your ELISA kit, it is best practice to validate that the standard curve generated is parallel to that of the kit’s recommended standard, ensuring accurate quantification.

Best practices:

• Use recombinant M-CSF of high purity (>95%) and confirmed bioactivity.
• Prepare the standard curve in the same matrix as your samples when possible to minimize matrix effects.
• Validate parallelism between your recombinant standard and the kit standard if using a different source.

In summary, recombinant mouse M-CSF is widely accepted and validated as a standard for ELISA quantification, provided it is appropriately prepared and validated for your specific assay system.

Recombinant Mouse M-CSF has been validated in published research for a range of applications, primarily related to the differentiation, proliferation, and functional analysis of monocytes, macrophages, and osteoclasts.

Key validated applications include:

• Functional Assays: Used to assess biological activity, such as stimulating proliferation of the M-NFS-60 cell line, a standard bioassay for M-CSF activity.
• Cell Differentiation: Widely used to generate bone marrow-derived macrophages (BMDMs) from mouse progenitor cells and to induce osteoclast differentiation from precursors.
• Proliferation and Survival Studies: Supports proliferation and survival of monocytes, macrophages, and their progenitors in vitro.
• Macrophage Polarization: Induces differentiation toward anti-inflammatory (M2-like) macrophages, relevant for immunological and inflammation studies.
• Phagocytosis and Functional Activation: Enhances phagocytic activity, cytokine production, and other effector functions in monocytes and macrophages.
• ELISA, Western Blot, Immunohistochemistry: Validated as a standard or control in immunoassays for detection and quantification of M-CSF or related markers.
• Disease Modeling: Used in preclinical models to study roles in neuroinflammation, bone resorption, atherosclerosis, cancer, and tissue repair, including microglial activation and remyelination in the CNS.
• Therapeutic Research: Investigated in studies targeting M-CSF/CSF1R signaling for potential therapies in cancer, autoimmune, and inflammatory diseases.

Summary Table of Validated Applications

These applications are supported by both product validation data and numerous peer-reviewed studies, confirming the broad utility of recombinant mouse M-CSF in immunology, cell biology, and disease modeling research.

To reconstitute and prepare Recombinant Mouse M-CSF protein for cell culture experiments, dissolve the lyophilized protein in sterile water or buffer to a concentration between 0.1–1.0 mg/mL. For most applications, a concentration of 0.1 mg/mL is recommended.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to ensure all powder is at the bottom.
• Add sterile distilled water or PBS (pH 7.2–8.0) to achieve the desired concentration (e.g., for 100 µg, add 1 mL for 0.1 mg/mL or 100 µL for 1 mg/mL).
• Gently pipet and wash down the sides of the vial to fully dissolve the protein. Do not vortex; gentle mixing is preferred to avoid denaturation.
• If the protein appears as a film, ensure it is fully dissolved by gentle mixing.
• For long-term storage, aliquot the reconstituted solution and store at –20°C or –80°C. Add a carrier protein (e.g., 0.1% BSA or HSA) to prevent adsorption and loss of activity during freezing.
• For short-term use, store at 2–8°C for up to one week.
• Avoid repeated freeze-thaw cycles to maintain protein integrity.

Preparation for cell culture:

• Before adding to cells, dilute the stock solution to the desired working concentration in cell culture medium. Typical working concentrations for bioassays range from 0.5–3 ng/mL, but optimal dosing should be determined empirically for your specific cell type and assay.
• Ensure the final solution is sterile and compatible with your cell culture system.

Additional notes:

• If the formulation contains only buffer and no carrier protein, adding 0.1% BSA or HSA after reconstitution is strongly recommended for stability, especially for storage or repeated use.
• Always check the manufacturer’s certificate of analysis for endotoxin levels and purity, as high endotoxin can affect cell culture results.

This protocol ensures maximal recovery and biological activity of recombinant Mouse M-CSF for cell culture applications.