Recombinant Human CCL23

Recombinant Human CCL23 is used in research applications to study immune cell recruitment, immune modulation, and tumor microenvironment dynamics due to its well-characterized chemotactic and immunoregulatory properties.

Key scientific reasons to use Recombinant Human CCL23:

• Chemotactic Activity: CCL23 is highly chemotactic for resting T cells, monocytes, and to a lesser extent, neutrophils. It is commonly used to investigate mechanisms of immune cell migration and trafficking in vitro and in vivo.
• Immune Modulation: CCL23 can induce T cell exhaustion markers (e.g., CTLA-4, PD-1, TIGIT, TIM-3, LAG-3) in CD8+ T cells, making it valuable for studies on immune checkpoint regulation and tumor immunology.
• Tumor Microenvironment Research: CCL23 influences the tumor microenvironment (TME) by promoting immunosuppression in certain cancers (e.g., ovarian cancer) or enhancing antitumor immunity in others (e.g., hepatocellular carcinoma) through modulation of ER stress and cytokine profiles.
• Functional Assays: Recombinant CCL23 is validated for use in functional assays, ELISA, blocking assays, and Western blot, allowing for quantitative and qualitative analysis of its biological effects.
• Angiogenesis and Inflammation: CCL23 may play a direct role in angiogenesis and is involved in both acute and chronic inflammatory responses, making it relevant for vascular biology and inflammation studies.
• Pathway Analysis: CCL23 interacts with CCR1 and participates in chemokine signaling pathways, providing a model for receptor-ligand studies and downstream signaling investigations.

Technical advantages of recombinant protein:

• High purity and low endotoxin: Recombinant preparations typically offer >95–97% purity and <1 EU/µg endotoxin, ensuring reproducibility and minimizing confounding effects in cell-based assays.
• Defined activity: Recombinant CCL23 is biologically active and can be used at defined concentrations to elicit dose-dependent responses in immune cells.
• Versatility: Suitable for a range of applications including SDS-PAGE, HPLC, ELISA, and functional cell migration assays.

Typical research applications:

• Immune cell migration assays
• Tumor immunology and checkpoint studies
• Inflammation and angiogenesis models
• Chemokine signaling pathway analysis
• Functional blocking and neutralization assays

Using recombinant human CCL23 enables precise, reproducible, and mechanistically informative experiments in immunology, oncology, and cell biology.

Yes, you can use recombinant human CCL23 as a standard for quantification or calibration in your ELISA assays, provided it is properly characterized and its concentration is accurately determined.

Supporting details:

• ELISA standards are commonly recombinant proteins: Most commercial CCL23 ELISA kits use recombinant human CCL23 or synthetic peptides as their standard for generating the calibration curve. This is standard practice for quantitative ELISA assays.
• Quantification requires a standard curve: For accurate quantification, a standard curve must be prepared using known concentrations of the target protein, which can be a purified recombinant protein. The recombinant CCL23 should be reconstituted and diluted according to best practices to cover the expected concentration range in your samples.
• Kit compatibility: Many ELISA kits are validated to detect both natural and recombinant forms of human CCL23, indicating that recombinant protein is suitable for use as a standard.
• Critical considerations:
  • Ensure the recombinant CCL23 is of high purity and its concentration is precisely known (e.g., determined by absorbance at 280 nm, BCA, or Bradford assay).
  • The recombinant protein should match the sequence and post-translational modifications (if relevant) of the endogenous CCL23 detected by your assay, as differences can affect antibody recognition and quantification accuracy.
  • Prepare the standard curve in the same buffer/matrix as your samples to minimize matrix effects.

Best practices:

• Prepare a serial dilution series of the recombinant CCL23 to generate a standard curve spanning the expected sample concentrations.
• Validate the linearity and sensitivity of your standard curve within the dynamic range of your assay.
• If using a custom or non-kit recombinant CCL23, confirm its compatibility with your specific ELISA antibodies and detection system.

In summary, recombinant human CCL23 is appropriate and widely used as a standard for ELISA quantification, provided it is well-characterized and handled according to established protocols.

Recombinant Human CCL23 has been validated for several applications in published research, including functional assays, chemotaxis studies, ELISA, Western blot, and blocking assays.

Key validated applications and research uses include:

• Functional Assays: CCL23 is widely used in chemotaxis assays to demonstrate its ability to attract immune cells such as monocytes, resting T lymphocytes, and neutrophils. It has also been used to study its effects on endothelial cell migration, differentiation, and neovascularization in vivo (e.g., chick chorioallantoic membrane assay).
• ELISA: Recombinant CCL23 is used as a standard or analyte in ELISA kits for quantifying CCL23 levels in biological samples.
• Western Blot: It is validated for use in Western blotting to detect CCL23 protein or to assess downstream signaling events (e.g., phosphorylation of kinases such as GSK3β in T cells).
• Blocking Assays: CCL23 has been used in blocking assays to study receptor-ligand interactions, particularly with its receptor CCR1.
• Flow Cytometry: In vitro studies have used recombinant CCL23 to treat immune cells (e.g., CD8+ T cells), followed by flow cytometry to assess upregulation of exhaustion markers (CTLA-4, PD-1, TIGIT, TIM-3, LAG-3).
• Cellular Signaling Studies: CCL23 has been used to investigate intracellular signaling pathways, such as IP3 production and kinase phosphorylation in vascular smooth muscle cells and T cells.
• Colony Formation Assays: It has been used to assess inhibition of myeloid progenitor cell proliferation.

Summary Table of Validated Applications

These applications are supported by both product validation data and peer-reviewed research, demonstrating the utility of recombinant human CCL23 in immunology, cancer biology, vascular biology, and cell signaling studies.

To reconstitute and prepare Recombinant Human CCL23 protein for cell culture experiments, dissolve the lyophilized protein in sterile water or an aqueous buffer containing 0.1% BSA (bovine serum albumin) to your desired concentration. This approach helps maintain protein stability and prevents adsorption to tube surfaces.

Step-by-step protocol:

1. Centrifuge the vial briefly to ensure all lyophilized powder is at the bottom before opening.
2. Reconstitution:
   • Add sterile, endotoxin-free water or 0.1% BSA in PBS (phosphate-buffered saline) to the vial.
   • The recommended concentration for reconstitution is typically 0.1–1.0 mg/mL.
   • Gently pipette up and down or swirl to dissolve. Avoid vigorous vortexing to prevent protein denaturation.
3. Optional: For enhanced stability, especially for long-term storage or repeated freeze-thaw cycles, add 5–50% glycerol to the reconstituted solution.
4. Aliquot the solution to avoid repeated freeze-thaw cycles, which can degrade the protein.
5. Storage:
   • Store aliquots at −20°C or below for long-term use (up to several months).
   • For short-term use (within 1 week), store at 4°C.
6. Working concentration: For cell-based assays, CCL23 is typically used at 10–50 ng/mL, but optimal concentrations should be determined empirically for your specific cell type and assay.

Additional notes:

• Always use sterile technique to avoid contamination.
• If the protein is to be used in sensitive cell culture systems, ensure the final buffer is compatible with your cells (e.g., isotonic, physiological pH).
• If the protein was supplied in a buffer containing acetonitrile or trifluoroacetic acid, consider buffer exchange or dialysis into PBS or cell culture medium before use.

Summary Table:

This protocol ensures the protein remains stable and bioactive for cell culture applications.