Recombinant Mouse MIP-1β

Recombinant Mouse MIP-1β (CCL4) is a chemokine widely used in research to study immune cell recruitment, inflammation, and disease mechanisms due to its potent ability to chemoattract and activate various leukocyte populations.

Key scientific reasons to use Recombinant Mouse MIP-1β in research applications:

• Immune Cell Recruitment: MIP-1β is a strong chemoattractant for CD4+ T cells, monocytes, and NK cells, making it essential for studying leukocyte migration and trafficking in models of inflammation, infection, and cancer.

• Inflammatory Response Modulation: It induces the release of proinflammatory cytokines, mast cell degranulation, and neutrophil activation (including superoxide production), allowing detailed analysis of acute and chronic inflammatory processes.

• Disease Modeling: MIP-1β is implicated in various disease models, such as:

  • Autoimmunity: Suppresses islet beta-cell inflammatory responses, relevant for diabetes research.
  • Cancer: Regulates tumoricidal monocyte activity and is involved in tumor microenvironment studies.
  • Infection and Sepsis: Used in models of pneumonia and sepsis to study immune cell dynamics and host defense.
  • Tissue Engineering: Contributes to immune privilege and regeneration in cartilage tissue engineering.

• Receptor Specificity: MIP-1β primarily signals through CCR5, and to a lesser extent CCR1 and CCR2, enabling mechanistic studies of chemokine-receptor interactions and downstream signaling.

• Bioassay and Functional Studies: Recombinant MIP-1β is validated for use in bioassays, including chemotaxis assays, cytokine release studies, and functional immune cell activation protocols.

• Species Compatibility: Recombinant mouse MIP-1β is specifically suited for murine models, ensuring physiological relevance and compatibility with mouse immune systems.

Best Practices:

• Use carrier-free preparations for sensitive bioassays to avoid interference.
• Avoid repeated freeze/thaw cycles to maintain protein activity.
• Validate activity by chemoattraction assays (e.g., Baf3-hCCR5 cells).

Summary of Applications:

• Immunology (cell migration, cytokine profiling)
• Inflammation (acute/chronic models)
• Cancer (tumor microenvironment, metastasis)
• Autoimmunity (diabetes, lupus nephritis)
• Infectious disease (sepsis, pneumonia)
• Tissue engineering (immune privilege, regeneration)

In conclusion, recombinant mouse MIP-1β is a versatile tool for dissecting immune mechanisms, modeling disease, and testing therapeutic interventions in murine systems due to its well-characterized biological activities and receptor interactions.

Yes, recombinant mouse MIP-1β can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated for your specific assay system. Recombinant proteins are commonly used as standards in quantitative ELISA protocols, as they allow for the generation of a standard curve against which unknown sample concentrations can be measured.

Key considerations and supporting details:

• Assay Compatibility: The recombinant mouse MIP-1β must be recognized by the capture and detection antibodies used in your ELISA. Commercial mouse MIP-1β ELISA kits are typically calibrated using recombinant mouse MIP-1β, and validation data show that these standards yield accurate and parallel dose-response curves when compared to natural protein.

• Standard Preparation: The recombinant protein should be highly purified and its concentration accurately determined. Standards are usually prepared by serial dilution in the same buffer or matrix as your samples to minimize matrix effects.

• Validation: It is important to confirm that the standard curve generated with your recombinant mouse MIP-1β is linear and covers the expected concentration range of your samples. Validation studies have shown that recombinant cytokine standards, including mouse MIP-1β, provide reliable quantification when used in ELISA, with high correlation to natural protein measurements.

• Documentation: Always follow the manufacturer’s instructions for reconstitution, dilution, and storage of the recombinant standard, as these can affect assay performance.

• Species Specificity: Ensure that the recombinant standard matches the species and isoform of the analyte you intend to measure (i.e., mouse MIP-1β for mouse samples).

Summary Table: Use of Recombinant Mouse MIP-1β as ELISA Standard

In conclusion, recombinant mouse MIP-1β is suitable as a standard for ELISA quantification, provided it is validated for your assay and handled according to best practices.

Recombinant Mouse MIP-1β (CCL4) has been validated for use in bioassays, cell culture, in vivo studies, ELISA, functional assays, and SDS-PAGE in published research.

Key validated applications include:

• Bioassay: Used to assess chemotactic activity and cellular responses, such as chemoattraction of Baf3-hCCR5 cells and modulation of immune cell behavior in various disease models including glioma, lupus nephritis, cartilage tissue engineering, and tumor angiogenesis.
• Cell Culture: Applied to study immune cell activation, cytokine signaling, and interactions in vitro, particularly in T cell and macrophage research.
• In Vivo Studies: Utilized in animal models to investigate inflammatory responses, disease progression (e.g., sepsis, acute lung injury, fever induction), and immune cell infiltration.
• ELISA (Enzyme-Linked Immunosorbent Assay): Used as a standard or analyte for quantifying MIP-1β levels in biological samples.
• Functional Assay: Employed to measure biological activity, such as chemotaxis and cytokine induction.
• SDS-PAGE: Used for protein characterization and purity assessment.
• Western Blot: Validated for detection and quantification of MIP-1β protein in samples.

Representative published research applications:

• Asthma and glioma formation: Investigating T cell-mediated inhibition of microglia in mouse models.
• Sepsis-induced pneumonia: Studying immune modulation in vivo.
• Tumor angiogenesis: Assessing myeloid cell-dependent mechanisms in cancer models.
• Lupus nephritis: Exploring inflammatory cell infiltration in kidney disease.
• Cartilage tissue engineering: Enhancing regeneration via immune privilege mechanisms.
• Acute lung injury: Evaluating chemokine roles in inflammatory lung disease.
• Fever induction: Studying pyrogenic responses and chemokine receptor interactions in vivo.

Additional validated uses:

• Blocking Assay: Used to assess inhibition of receptor-ligand interactions.
• Therapeutic target studies: Investigated for its role in chronic inflammation, autoimmune diseases, and as a potential therapeutic target.

These applications are supported by multiple peer-reviewed studies and product validation data, demonstrating the versatility of recombinant mouse MIP-1β in immunology, inflammation, and disease modeling research.

To reconstitute and prepare Recombinant Mouse MIP-1β (CCL4) protein for cell culture experiments, dissolve the lyophilized protein at a concentration of 100 μg/mL in sterile PBS containing at least 0.1% human or bovine serum albumin (BSA or HSA) as a carrier protein. If the formulation is carrier-free or contains trehalose, reconstitution in sterile PBS alone is sufficient.

Step-by-step protocol:

• Equilibrate the vial and reconstitution buffer to room temperature before opening.
• Centrifuge the vial briefly to collect all lyophilized material at the bottom.
• Add buffer: For most applications, use sterile PBS with 0.1–1% BSA or HSA. For carrier-free formulations, PBS alone is acceptable.
• Gently pipette to wash down the sides of the vial and ensure complete dissolution of the protein.
• Incubate for 15–30 minutes at room temperature with gentle agitation. If undissolved flakes remain, continue mixing for up to 2 hours.
• Aliquot the stock solution to avoid repeated freeze-thaw cycles.
• Storage: Store aliquots at −20 °C or colder. For short-term use (up to one week), aliquots can be kept at 2–8 °C.
• Avoid repeated freeze/thaw cycles to maintain protein integrity.

Preparation for cell culture:

• Dilute the stock solution to the desired working concentration using cell culture medium immediately before use.
• Ensure the final concentration of carrier protein (BSA/HSA) does not interfere with your assay or cell type.

Additional notes:

• Always consult the specific product datasheet for formulation details, as some preparations may contain additives (e.g., acetonitrile, TFA, trehalose) that could affect reconstitution or downstream applications.
• For bioassays, confirm that the recombinant protein is tested for biological activity in cell-based systems.
• If using for ELISA or other analytical assays, follow the recommended reconstitution buffer and protocol for that application, as requirements may differ.

Summary of key points:

• Concentration: 100 μg/mL
• Buffer: Sterile PBS + 0.1–1% BSA/HSA (unless carrier-free)
• Aliquot and store: −20 °C or colder
• Avoid freeze/thaw cycles
• Dilute in cell culture medium for experiments

These steps will ensure optimal solubility, stability, and biological activity of recombinant Mouse MIP-1β for cell culture experiments.