Recombinant Mouse MIP-1α

Recombinant Mouse MIP-1α (also known as CCL3) is widely used in research to study immune cell recruitment, inflammation, and immune modulation, particularly in mouse models. Its applications are grounded in its well-characterized biological activities and its role as a key chemokine in both innate and adaptive immunity.

Key reasons to use recombinant Mouse MIP-1α in research:

• Chemoattractant Activity: MIP-1α is a potent chemoattractant for monocytes, granulocytes, and other immune cells, facilitating studies on cell migration, recruitment, and trafficking in vitro and in vivo.
• Immune Modulation: It modulates immune responses by binding to chemokine receptors CCR1, CCR4, and CCR5, leading to the recruitment and activation of various leukocyte subsets, including granulocytes and monocytes.
• Inflammation and Disease Models: MIP-1α is critical in models of inflammation, infection, wound healing, and autoimmune diseases, making it valuable for dissecting mechanisms of immune cell infiltration and inflammatory signaling.
• Adjuvant Effects: When co-administered with vaccines or antigens, MIP-1α can enhance antigen-specific CD8+ T cell responses, promote Th1 polarization, and increase resistance to viral challenges, supporting its use as an immuno-adjuvant in vaccine research.
• Tumor Immunology: MIP-1α can induce tumoricidal monocyte activity and regulate tumor growth, making it relevant for cancer immunology studies.
• Neuroscience: It acts as a neuromodulator in the hippocampus, influencing synaptic plasticity and cognitive functions, thus being useful in neuroinflammation and neurodegeneration research.
• Standardization and Reproducibility: Recombinant proteins provide consistent, defined activity and purity, which is essential for reproducible experimental results.

Typical applications include:

• Chemotaxis assays to study immune cell migration.
• In vivo models of inflammation, infection, wound healing, and autoimmune disease.
• Immunization protocols to enhance cellular immune responses.
• Functional studies of chemokine receptor signaling.
• Investigation of neuroimmune interactions.

In summary, recombinant Mouse MIP-1α is a versatile tool for dissecting immune mechanisms, modeling disease processes, and enhancing immunological interventions in mouse-based research.

You can use recombinant Mouse MIP-1α as a standard for quantification or calibration in ELISA assays, provided it is of high purity, its concentration is accurately known, and it is compatible with your assay system.

Key considerations and supporting details:

• Recombinant proteins are commonly used as ELISA standards because they provide a consistent, well-characterized source of antigen for generating standard curves. Many commercial ELISA kits for Mouse MIP-1α (also known as CCL3) use recombinant protein as the standard, and these kits demonstrate that natural and recombinant forms yield parallel standard curves, indicating equivalence for quantification.

• Purity and quantification: The recombinant protein should be highly purified (typically >95%) and its concentration should be determined accurately, ideally by absorbance at 280 nm or amino acid analysis. Impurities or inaccurate quantification can introduce error into your standard curve.

• Buffer compatibility: Ensure the recombinant protein is reconstituted or diluted in a buffer compatible with your ELISA system, often matching the matrix of your samples (e.g., assay diluent or sample buffer).

• Validation: If you are not using the standard provided with a commercial kit, you should validate that your recombinant standard produces a standard curve that is parallel to the curve generated by the kit standard or by natural MIP-1α in your sample matrix. This ensures that the antibody pair in your ELISA recognizes the recombinant form equivalently.

• Bioassay vs. ELISA standards: Some recombinant proteins are specifically labeled for use as ELISA standards and not for functional bioassays, as they may not be tested for biological activity. For quantification/calibration in ELISA, only immunoreactivity is required.

• Lot-to-lot variation: If you prepare your own standards from recombinant protein, be aware of potential lot-to-lot variation in protein content or activity. Always calibrate each new lot.

Summary Table: Recombinant Mouse MIP-1α as ELISA Standard

If you follow these guidelines, recombinant Mouse MIP-1α is suitable as a standard for ELISA quantification and calibration. Always consult your specific ELISA protocol for any additional requirements.

Recombinant Mouse MIP-1α (CCL3) has been validated in published research for a range of applications, primarily related to its chemotactic and immunomodulatory functions. The most common validated applications include:

• Chemotaxis assays: Used to demonstrate the ability of MIP-1α to chemoattract various cell types, such as human monocytes, mouse BaF3 cells transfected with human CCR5, B cells, eosinophils, and neutrophils.
• Bioassays: Functional assays measuring cellular activation, adhesion, and migration in response to MIP-1α, often using whole cells or primary immune cells.
• In vivo studies: Used to study leukocyte recruitment, inflammation, and immune responses in mouse models, including wound healing, infection, and autoimmune disease models.
• Western blot: Detection and quantification of MIP-1α protein in tissue or cell lysates.
• ELISA: Quantitative measurement of MIP-1α in biological samples, such as serum or tissue culture supernatants.
• Functional inhibition and receptor binding studies: Used to characterize interactions with chemokine receptors (CCR1, CCR5) and to study receptor pharmacology.
• Mass spectrometry, HPLC, SDS-PAGE: For protein characterization, purity assessment, and quality control.

Published research examples:

• Chemotaxis and immune cell recruitment: MIP-1α has been used to study neutrophil and monocyte migration in vitro and in vivo, including in models of wound healing, sepsis, and inflammation.
• Stem cell regulation: MIP-1α has been shown to inhibit hematopoietic stem cell proliferation in vitro and in vivo.
• Cancer and immunotherapy: Used in studies of dendritic cell vaccines and tumor microenvironment modulation.
• Neuroinflammation: Applied in mouse models of Alzheimer’s disease to study astrocyte activation and phagocytosis.

Summary Table of Validated Applications

Key points:

• The most robustly validated applications are chemotaxis and functional bioassays, both in vitro and in vivo.
• MIP-1α is widely used to study immune cell migration, inflammation, and hematopoietic regulation.
• Additional uses include protein detection (Western blot, ELISA) and receptor pharmacology.

If you need protocols or more specific details for any application, please specify the context or cell type of interest.

To reconstitute and prepare Recombinant Mouse MIP-1α (CCL3) protein for cell culture experiments, dissolve the lyophilized protein at a concentration of 100 μg/mL in sterile PBS. For optimal stability and to prevent adsorption, add 0.1%–1% carrier protein such as bovine serum albumin (BSA) or human serum albumin (HSA) if the protein is not already supplied with a carrier.

Detailed protocol:

• Centrifuge the vial briefly to collect all lyophilized material at the bottom before opening.
• Add sterile PBS (phosphate-buffered saline) to achieve a final concentration of 100 μg/mL. For example, add 100 μL PBS to 100 μg protein.
• Include carrier protein: If the product is carrier-free, add 0.1%–1% BSA or HSA to the PBS before reconstitution to minimize protein loss due to adsorption.
• Gently mix by pipetting or slow vortexing. Avoid vigorous agitation or foaming.
• Allow the protein to dissolve at room temperature for 10–30 minutes.
• Aliquot the solution to avoid repeated freeze-thaw cycles, which can reduce activity.
• Storage: Store aliquots at −20 °C or below for long-term storage. For short-term use (up to one week), store at 2–8 °C.

Additional notes:

• If the protein is supplied in a low pH buffer (e.g., glycine, pH 3.0), you may dilute further in cell culture medium or PBS with carrier protein to adjust the pH before use in cell assays.
• Always consult the product-specific datasheet for any unique instructions, as formulations may vary.
• Avoid repeated freeze/thaw cycles to maintain protein integrity and activity.

Typical working concentrations for cell assays range from 0.1–10 ng/mL, but optimal dosing should be determined empirically for your specific application.

This protocol ensures maximal recovery and biological activity of recombinant MIP-1α for cell culture experiments.