Recombinant Mouse Macrophage Inflammatory Protein 1 Gamma (MIP-1γ)

Recombinant Mouse Macrophage Inflammatory Protein 1 Gamma (MIP-1γ, also known as CCL9/10) is a chemokine with key roles in immune cell recruitment, osteoclast biology, and inflammatory signaling, making it a valuable tool for research in immunology, inflammation, and bone metabolism.

Key reasons to use recombinant MIP-1γ in research applications:

• Chemotactic Activity: MIP-1γ is a potent chemoattractant for various immune cells, including neutrophils and monocytes, enabling the study of cell migration, trafficking, and recruitment in vitro and in vivo.
• Osteoclast Differentiation and Survival: MIP-1γ plays an important role in the differentiation and survival of osteoclasts, making it relevant for research on bone remodeling, osteoporosis, and related pathologies.
• Modeling Inflammatory Responses: As a member of the MIP-1 chemokine family, MIP-1γ is involved in the regulation of inflammatory and immune responses, allowing researchers to model and dissect mechanisms underlying inflammation, autoimmune diseases, and host defense.
• Receptor-Ligand Studies: MIP-1γ acts via G-protein-coupled receptors (notably CCR1, CCR3, and CCR5), providing a system to study chemokine-receptor interactions, signaling pathways, and the effects of receptor antagonists.
• Cellular Source and Expression: MIP-1γ is constitutively expressed in macrophages, dendritic cells, and myeloid cell lines, making recombinant protein essential for controlled, reproducible experiments where endogenous expression may be variable or insufficient.
• Functional Assays: Recombinant MIP-1γ can be used in chemotaxis assays, cell differentiation protocols, cytokine release studies, and in vivo models to assess its biological effects under defined conditions.

Additional context:

• MIP-1γ is part of the CC chemokine subfamily, which includes other well-studied members like MIP-1α (CCL3) and MIP-1β (CCL4), all of which are important in immune regulation and inflammation.
• The use of recombinant proteins ensures batch-to-batch consistency and eliminates variability from endogenous sources, which is critical for reproducibility in experimental research.

Summary of applications:

• Immunology: Study of leukocyte migration, immune cell activation, and chemokine signaling.
• Inflammation: Modeling acute and chronic inflammatory responses.
• Bone Biology: Investigation of osteoclastogenesis and bone resorption mechanisms.
• Drug Discovery: Screening and validation of chemokine receptor antagonists or modulators.

Using recombinant MIP-1γ allows for precise control over experimental conditions and facilitates mechanistic studies in mouse models and cell-based systems.

Yes, recombinant Mouse Macrophage Inflammatory Protein 1 Gamma (MIP-1γ) can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and its concentration is accurately determined. This is a common practice in ELISA development and commercial kits, where recombinant proteins serve as standards to generate calibration curves for quantifying the target analyte in biological samples.

Essential context and best practices:

• Standard Curve Generation: ELISA kits for mouse MIP-1γ typically use recombinant MIP-1γ as the standard to generate a standard curve, which is then used to interpolate the concentration of MIP-1γ in unknown samples.
• Purity and Quantification: The recombinant protein used as a standard should be highly purified and its concentration precisely quantified, ideally by absorbance at 280 nm using a known extinction coefficient or by amino acid analysis. Impurities or inaccurate quantification can lead to errors in assay calibration.
• Buffer Compatibility: The recombinant standard should be reconstituted and diluted in the same buffer as the samples and assay diluent to minimize matrix effects and ensure accurate quantification.
• Validation: If you are developing your own ELISA or using a non-kit recombinant standard, it is important to validate that the recombinant MIP-1γ is recognized equivalently by the capture and detection antibodies used in your assay. This ensures that the standard curve accurately reflects the detection of endogenous MIP-1γ in your samples.
• Species Specificity: Ensure that the recombinant protein matches the species and isoform of MIP-1γ you intend to measure, as cross-reactivity or sequence differences can affect antibody binding and quantification accuracy.

Additional relevant information:

• Commercial ELISA kits for mouse MIP-1γ (also known as CCL9/CCL10) routinely use recombinant mouse MIP-1γ as the standard, with typical detection ranges from low pg/mL to several hundred pg/mL.
• If using a recombinant standard from a different source than the kit manufacturer, it is advisable to compare its performance with the kit standard to ensure consistent results.

In summary, using recombinant mouse MIP-1γ as a standard is scientifically appropriate and widely practiced in ELISA quantification, provided the above considerations are addressed for assay accuracy and reproducibility.

Recombinant Mouse Macrophage Inflammatory Protein 1 Gamma (MIP-1γ) has been validated in published research primarily for applications related to immune cell migration, T cell polarization, and osteoclast biology.

Key validated applications include:

• Chemotaxis assays: MIP-1γ has been shown to mediate the migration of CD4⁺ and CD8⁺ T cells, indicating its use in chemotaxis and cell recruitment studies.
• T cell polarization studies: It is selectively produced by Th1 cells (not Th2), and recombinant MIP-1γ is used to investigate Th1/Th2 differentiation and cytokine profiles in vitro and in vivo.
• Osteoclast differentiation and survival: MIP-1γ plays a role in the differentiation and survival of osteoclasts, supporting its use in bone biology and osteoclastogenesis assays.
• Functional immunology assays: Recombinant MIP-1γ is used to study its effects on immune cell activation, cytokine production, and potential contributions to parasite clearance and inflammatory responses.
• Gene expression and protein detection: MIP-1γ mRNA and protein have been detected in dendritic cells and Langerhans cells, validating its use in gene expression profiling and protein quantification experiments.

Additional context:

• MIP-1γ is a CC chemokine produced by various immune cells, including macrophages, dendritic cells, and lymphocytes.
• Its chemotactic properties are leveraged to study immune cell trafficking in models of infection, inflammation, and tissue injury.
• The protein is also implicated in the regulation of cellular immune responses, including recruitment of cytotoxic T cells to sites of inflammation or infection.

These applications are supported by published studies using recombinant mouse MIP-1γ in both in vitro and in vivo experimental systems, particularly in the context of immunology, inflammation, and bone metabolism research.

To reconstitute and prepare Recombinant Mouse Macrophage Inflammatory Protein 1 Gamma (MIP-1γ) for cell culture experiments, follow these steps:

• Centrifuge the vial briefly before opening to ensure all lyophilized protein is at the bottom.
• Reconstitute the protein in sterile, distilled water to a final concentration of 0.1–1.0 mg/mL.
• Gently mix to dissolve the protein completely. Avoid vigorous vortexing to prevent protein denaturation.
• For working concentrations, make further dilutions in an aqueous buffer (such as PBS or cell culture medium) supplemented with a carrier protein like 0.1–1.0% BSA to stabilize the protein and minimize adsorption to plastic surfaces.
• Aliquot the reconstituted protein to avoid repeated freeze-thaw cycles, which can reduce activity.
• Storage:
  • Before reconstitution, store the lyophilized protein at –20 °C.
  • After reconstitution, store aliquots at –20 °C or below for long-term storage, or at 2–8 °C for short-term use (up to 1 month).

Additional notes for cell culture use:

• Always use sterile technique to prevent contamination.
• If using in functional assays (e.g., chemotaxis), confirm the biological activity of the reconstituted protein with a pilot experiment.
• Prepare fresh dilutions for each experiment to ensure reproducibility.

Summary protocol:

1. Briefly centrifuge vial.
2. Add sterile distilled water to achieve 0.1–1.0 mg/mL.
3. Mix gently until fully dissolved.
4. Dilute to working concentration in buffer with 0.1–1.0% BSA.
5. Aliquot and store appropriately.

These steps will ensure optimal solubility, stability, and activity of recombinant MIP-1γ for cell culture applications.