Recombinant Human MIF

Recombinant Human Macrophage Migration Inhibitory Factor (MIF) is a valuable tool for a wide range of research applications due to its well-characterized biological activities and relevance in immune regulation, inflammation, and disease pathogenesis. Here are several key reasons why you should consider using Recombinant Human MIF in your research:

1. Well-Defined Biological Functions

Recombinant Human MIF is a non-glycosylated polypeptide (typically ~12.5 kDa) that retains the full biological activity of native MIF. It is known to:

• Stimulate the production of pro-inflammatory cytokines such as TNF-α, IL-1, and IL-8.
• Induce nitric oxide (NO) release in activated macrophages.
• Regulate the migration and activation of immune cells, including macrophages and T cells.
• Modulate matrix metalloproteinase (MMP) expression, influencing tissue remodeling and fibrosis.

2. Versatility in Experimental Models

Recombinant Human MIF is suitable for use in:

• In vitro assays: Cell culture studies with macrophages, fibroblasts, endothelial cells, and other cell types to investigate inflammatory responses, cell migration, and signaling pathways.
• In vivo studies: Animal models of inflammation, infection, autoimmune diseases, and cancer to assess the role of MIF in disease progression and therapeutic interventions.
• Bioassays and functional studies: Evaluating the effects of MIF on cytokine production, cell survival, and immune modulation.

3. High Purity and Consistency

Commercially available Recombinant Human MIF is typically produced in E. coli and purified to >95–97% purity. This ensures reproducibility and reliability in experimental results, minimizing batch-to-batch variability.

4. Carrier-Free Options

Many suppliers offer carrier-free versions of Recombinant Human MIF, which are ideal for sensitive applications such as surface plasmon resonance (SPR), ELISA, and other assays where the presence of carrier proteins (e.g., BSA) could interfere with results.

5. Relevance to Disease Mechanisms

MIF plays a critical role in various pathological conditions, including:

• Inflammatory diseases: Arthritis, colitis, septic shock.
• Cardiovascular diseases: Atherosclerosis, myocardial infarction.
• Cancer: Tumor microenvironment, immune evasion, and angiogenesis.
• Infectious diseases: Host defense against pathogens, viral persistence, and immune modulation.

Using Recombinant Human MIF allows researchers to dissect the molecular mechanisms underlying these diseases and evaluate potential therapeutic targets.

6. Compatibility with Antibody and Inhibitor Studies

Recombinant Human MIF is widely used as a positive control in immunoassays and for generating or validating MIF-specific antibodies and inhibitors. It is also essential for studying the effects of MIF-neutralizing agents or small molecule inhibitors in preclinical models.

7. Support for Translational Research

Given its involvement in multiple disease pathways, Recombinant Human MIF supports translational research aimed at developing new diagnostics, prognostics, and therapeutics targeting MIF or its downstream effectors.

Summary

Recombinant Human MIF is a robust, reliable, and versatile reagent that enables researchers to investigate the complex roles of MIF in immunity, inflammation, and disease. Its high purity, biological activity, and compatibility with diverse experimental systems make it an indispensable tool for advancing both basic and applied research in immunology, oncology, and infectious diseases.

Yes, recombinant human MIF can be used as a standard for quantification or calibration in ELISA assays, provided it is properly purified and its concentration is accurately determined. Recombinant MIF is commonly used as a standard in commercial ELISA kits and is recognized by antibodies that detect both recombinant and native human MIF.

Essential context and supporting details:

• ELISA Standard Requirements: For accurate quantification, the standard should be a purified protein with a known concentration. Recombinant human MIF, when properly prepared and quantified, fulfills this requirement and is routinely used in ELISA kits for calibration.

• Antibody Recognition: Most sandwich ELISA kits for MIF use antibodies that recognize both recombinant and natural forms of human MIF, ensuring that the standard curve generated with recombinant MIF is valid for quantifying endogenous MIF in biological samples.

• Validation and Linearity: Commercial kits report that recombinant human MIF yields linear standard curves and recovery rates comparable to those obtained with natural MIF, supporting its use for quantification.

• Formulation Considerations: Recombinant MIF is often supplied with carrier proteins (e.g., BSA) to enhance stability. For ELISA calibration, using the formulation recommended for standards (often with BSA) is advised unless the presence of carrier protein interferes with your assay.

• Preparation and Handling: Follow manufacturer or protocol-specific instructions for reconstitution, dilution, and storage of the recombinant MIF standard. Accurate pipetting and use of appropriate diluents are critical for reliable calibration.

Best practices:

• Use recombinant human MIF with verified purity and concentration.
• Prepare a standard curve covering the expected range of MIF concentrations in your samples (typically 0–1000 pg/mL, but may vary by kit and sample type).
• Validate recovery and linearity in your specific sample matrix if possible.
• Ensure compatibility of the recombinant MIF standard with the antibodies used in your ELISA.

Summary Table: Recombinant Human MIF as ELISA Standard

In conclusion, recombinant human MIF is suitable and widely accepted as a standard for ELISA quantification and calibration, provided it meets purity and quantification requirements and is compatible with your assay system.

Recombinant Human MIF has been validated for a broad range of applications in published research, primarily in bioassays, ELISA, in vivo studies, and surface plasmon resonance (SPR), as well as in functional and blocking assays.

Key validated applications include:

• Bioassays: Used to study MIF’s biological activity, such as its effects on cytokine production, immune cell activation, and signaling pathways in various cell types (e.g., cardiomyocytes, endothelial cells, macrophages, T cells).
• ELISA (Enzyme-Linked Immunosorbent Assay): Used as a standard or analyte to quantify MIF levels in biological samples and to assess MIF secretion in response to stimuli.
• In Vivo Studies: Applied in animal models to investigate MIF’s role in disease processes such as arthritis, myositis, and cardiovascular inflammation.
• Surface Plasmon Resonance (SPR): Used to characterize binding interactions between MIF and its ligands or inhibitors, including studies of MIF-receptor (CD74) interactions.
• Functional Assays: Employed to assess MIF’s biological functions, such as its pro-inflammatory effects, modulation of immune responses, and impact on cell migration and proliferation.
• Blocking Assays: Used to evaluate the efficacy of MIF inhibitors or neutralizing antibodies in blocking MIF activity.
• Western Blot: Occasionally used as a positive control or analyte for protein detection.
• Molecular and Cellular Mechanism Studies: Utilized in research on signaling pathways (e.g., ERK1/2, AMPK, AKT), cytokine release, and immune cell differentiation.

Representative published research applications:

• Investigating MIF’s role in cardiovascular inflammation and proinflammatory mediator production.
• Studying MIF-induced signaling in stem cell-derived exosome models and immune cell activation.
• Assessing MIF’s involvement in autoimmune diseases, such as rheumatoid arthritis, and its modulation of cytokine expression.
• Evaluating MIF as a target for small-molecule inhibitor development, including binding and functional inhibition studies.
• Using MIF as a biomarker in multiplex immunoassays for adipokine profiling.
• Exploring MIF’s effects in tumor microenvironments and its prognostic value in cancer research.

These applications demonstrate that recombinant human MIF is a versatile reagent for both basic and translational research in immunology, inflammation, cell signaling, and disease modeling.

To reconstitute and prepare Recombinant Human MIF 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). This formulation helps stabilize the protein and prevent adsorption to surfaces.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to ensure all lyophilized material is at the bottom.
• Add sterile PBS with 0.1% BSA to achieve the desired concentration (e.g., 100 μg/mL).
• Gently pipette the solution down the sides of the vial to fully dissolve the protein. Avoid vigorous mixing to prevent foaming and protein denaturation.
• Allow the protein to reconstitute for 15–30 minutes at room temperature with gentle agitation.
• Aliquot the solution if not using immediately, and store at −20 °C to −70 °C. Avoid repeated freeze-thaw cycles to maintain protein integrity.
• For cell culture, dilute the reconstituted stock solution in your culture medium to the desired working concentration, ensuring the final buffer is compatible with your cells.

Additional notes:

• If BSA is not suitable for your application, other carrier proteins (e.g., human serum albumin) may be used, but always verify compatibility with your assay and cell type.
• For sensitive applications, filter-sterilize the final solution using a 0.2 μm filter.
• Always consult the specific product datasheet for any additional formulation details or recommended diluents, as some recombinant proteins may require alternative buffers or additives.

Summary of best practices:

• Use sterile PBS + 0.1% BSA for reconstitution.
• Gentle mixing and room temperature incubation for complete dissolution.
• Aliquot and freeze unused stock to avoid freeze-thaw cycles.
• Dilute in cell culture medium for experimental use.

These steps will help ensure the stability and bioactivity of recombinant human MIF in cell culture experiments.