Recombinant Mouse CCL28

Recombinant mouse CCL28 offers multiple compelling advantages for research applications across diverse biological systems and disease models.

Immunomodulatory and Therapeutic Applications

Recombinant CCL28 demonstrates significant therapeutic potential in neuroinflammatory conditions. In spinal cord injury models, recombinant CCL28 promotes locomotor recovery by recruiting regulatory T cells (Treg) through the CCL28-CCR10 axis, creating an immunosuppressive microenvironment that reduces proinflammatory cytokine production and inhibits effector T cell proliferation. This mechanism establishes a direct causal link between CCL28 upregulation and functional recovery, making it valuable for studying neuroprotection and immune regulation in traumatic injury contexts.

Angiogenic and Vascular Applications

Recombinant CCL28 promotes angiogenesis across multiple tissue contexts. The protein enhances tube formation, migration, and proliferation of endothelial cells, with demonstrated efficacy in myocardial and hindlimb ischemia models. In ischemic tissues, recombinant CCL28 treatment improves blood flow recovery, increases vascular density, and reduces fibrosis, making it particularly useful for studying vascular regeneration and tissue repair mechanisms.

Antimicrobial and Antifungal Properties

Recombinant CCL28 exhibits potent antimicrobial activity against clinically relevant pathogens. The protein demonstrates rapid fungicidal activity against Candida albicans through direct membrane disruption mechanisms, showing superior efficacy compared to conventional antifungal agents. Both structured and unstructured recombinant CCL28 proteins induce negative Gaussian curvature on fungal membranes, causing solute leakage and membrane permeabilization. This makes recombinant CCL28 valuable for investigating novel antimicrobial mechanisms and developing alternative therapeutic strategies for opportunistic infections.

Mucosal Immunity and Adjuvant Functions

Recombinant CCL28 serves as a molecular adjuvant that enhances both systemic and mucosal immune responses. The protein regulates immunoglobulin A (IgA) antibody-secreting cell accumulation in mucosal tissues, controlling passive antibody transfer and mucosal immunity. This property makes it particularly useful for vaccine development research and mucosal immunology studies.

Experimental Advantages

The availability of recombinant mouse CCL28 enables controlled, dose-dependent studies without the variability associated with endogenous protein production. You can employ standardized concentrations in in vitro assays (transwell migration, tube formation, cell proliferation) and in vivo models with precise administration routes (intraspinal, topical, systemic). The protein's compatibility with neutralizing antibody approaches allows mechanistic studies through receptor blockade (CCR10, CCR3), enabling dissection of specific signaling pathways and validation of therapeutic targets.

Yes, recombinant mouse CCL28 can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to your assay system. Recombinant CCL28 is commonly used as the standard in commercial mouse CCL28 ELISA kits, where it is reconstituted and serially diluted to generate a standard curve for quantification.

Key considerations and supporting details:

• Recombinant CCL28 as Standard: Most commercial mouse CCL28 ELISA kits use recombinant mouse CCL28 as the standard protein for generating the calibration curve. This is standard practice in cytokine and chemokine ELISAs, as recombinant proteins provide a consistent and defined reference for quantification.

• Validation: It is essential that the recombinant CCL28 you use is of high purity, correctly folded, and biologically active (if required by your assay). The protein should be prepared in the same buffer as your samples or as recommended by your ELISA protocol to avoid matrix effects.

• Assay Compatibility: Ensure that the recombinant CCL28 is recognized by the capture and detection antibodies used in your ELISA. Most kits specify that their antibodies detect both natural and recombinant forms of mouse CCL28.

• Standard Preparation: Follow the ELISA kit or assay protocol for reconstitution and dilution of the recombinant standard. Typically, the standard is reconstituted in a specified buffer and serially diluted to cover the assay’s dynamic range.

• Limitations: If you are using a recombinant CCL28 from a source different from the one validated in your ELISA kit, you should verify that it produces a standard curve with expected sensitivity and linearity. Differences in glycosylation, folding, or sequence (e.g., tags or truncations) can affect antibody recognition and quantification accuracy.

• Documentation: Always document the source, lot, and concentration of your recombinant standard, and ideally validate it against a known standard if possible.

Summary Table: Use of Recombinant Mouse CCL28 as ELISA Standard

In conclusion, recombinant mouse CCL28 is widely accepted and used as a standard for ELISA quantification, but proper validation and protocol adherence are essential for accurate results.

Recombinant Mouse CCL28 has been validated for several key applications in published research, primarily in the fields of immunology, infection biology, angiogenesis, and functional protein assays.

Validated Applications:

• Immunology Research

  • Used to study chemotactic activity for immune cells such as CD4 and CD8 T-cells, eosinophils, and neutrophils via CCR3 and CCR10 signaling.
  • Investigated for its role in mucosal immunity and epithelial cell signaling.

• Antimicrobial and Infection Models

  • Demonstrated potent in vitro and in vivo antimicrobial activity, particularly against Candida albicans and Salmonella enterica, including membrane disruption and enhancement of neutrophil responses.
  • Used in mouse models of oropharyngeal candidiasis and bacterial infection to assess therapeutic potential and immune modulation.

• Angiogenesis and Tissue Repair

  • Validated in in vitro and in vivo angiogenesis assays, including Matrigel tube formation, chick chorioallantoic membrane (CAM) assays, and mouse models of myocardial and hindlimb ischemia.
  • Shown to promote neovascularization and improve blood flow recovery in ischemic tissues.

• Wound Healing Studies

  • Applied in diabetic mouse wound models to investigate the impact of CCL28 neutralization on wound healing, endothelial nitric oxide synthase (eNOS) expression, and vessel density.

• Functional Protein Assays

  • Used in functional assays (FuncS), blocking assays, and chemotaxis assays to characterize biological activity.
  • Validated as a standard in ELISA for quantification of CCL28 in biological samples.

• Protein Characterization

  • Analyzed by SDS-PAGE and HPLC for purity and structural validation.

Summary Table of Validated Applications

Key Insights:

• Recombinant Mouse CCL28 is widely used for both in vitro and in vivo studies, including cell migration, immune cell activation, antimicrobial activity, and tissue regeneration.
• Its activity is often mediated via CCR3 and CCR10 receptors, and it is a valuable tool for dissecting mucosal immune responses and angiogenic mechanisms.
• The protein is also validated for use as a standard in ELISA assays and for functional characterization in protein biochemistry.

If you require protocol details or specific experimental setups for any of these applications, please specify the area of interest.

To reconstitute and prepare Recombinant Mouse CCL28 protein for cell culture experiments, dissolve the lyophilized protein in sterile, endotoxin-free water or PBS to achieve a concentration between 0.1–1.0 mg/mL. For optimal stability and activity, follow these steps:

• Centrifuge the vial briefly before opening to ensure all powder is at the bottom.
• Add sterile water or PBS (check your product datasheet for specific recommendations; PBS is commonly used for chemokines) to reach your desired concentration, typically 0.1–1.0 mg/mL.
• Gently mix by pipetting up and down or swirling—do not vortex, as this may denature the protein.
• If recommended, add carrier protein (e.g., 0.1% BSA) to minimize adsorption and stabilize the protein, especially for low concentration working solutions.
• Aliquot the solution to avoid repeated freeze-thaw cycles.
• Store reconstituted protein at 2–8°C for short-term use (up to 1 month), or at −20°C to −80°C for long-term storage. Avoid repeated freeze-thaw cycles.

Preparation for cell culture:

• Before adding to cells, dilute the stock solution in your cell culture medium to the desired working concentration (often in the nanomolar range, e.g., 10–100 ng/mL, depending on your experimental design).
• Ensure the final buffer conditions (pH, osmolarity) are compatible with your cells and do not contain toxic additives.

Additional notes:

• If your application is sensitive to endotoxin, confirm the endotoxin level is <1 EU/μg, which is standard for recombinant chemokines.
• For enhanced stability, some protocols recommend adding 5–50% glycerol to the reconstituted solution for long-term storage.
• Always consult the specific product datasheet for any unique instructions regarding buffer composition or reconstitution volume.

This protocol ensures the recombinant CCL28 is properly solubilized, stable, and ready for use in cell culture assays.