Recombinant Human CD23

Recombinant Human CD23 is widely used in research applications because it is a critical molecule in immune regulation, especially in studies of B cell biology, IgE-mediated responses, and related diseases. Using the recombinant form provides a consistent, well-characterized, and species-specific tool for a variety of experimental needs.

Key reasons to use Recombinant Human CD23 in research:

• Immune Regulation Studies: CD23 (FcεRII) is the low-affinity receptor for IgE and plays a central role in regulating IgE-mediated immune responses, allergic reactions, and parasite immunity. It is involved in both upregulation and downregulation of IgE synthesis, making it essential for dissecting mechanisms of allergy and hypersensitivity.

• B Cell Function and Differentiation: CD23 is expressed on B cells and influences their growth, differentiation, and survival. It interacts with CD21 and other immune receptors, impacting B cell activation and antibody production.

• Disease Marker and Diagnostics: CD23 is a reliable immunophenotypic marker for distinguishing chronic lymphocytic leukemia (CLL, CD23-positive) from other B-cell malignancies. Its expression is also altered in various lymphomas and immune disorders.

• Functional Assays and Blocking Experiments: Recombinant CD23 can be used as a control or blocking agent in antibody validation, immunohistochemistry, immunocytochemistry, and western blotting to confirm antibody specificity. It is also used in ELISA and bioassays to study ligand binding and receptor function.

• Protein-Protein Interaction Studies: The recombinant protein enables detailed analysis of CD23’s interactions with IgE, CD21, MHC class II, and integrins, supporting research into immune complex formation and signaling pathways.

• Therapeutic Research: CD23 is a target for therapeutic antibody development in autoimmune and inflammatory diseases, as well as allergy and asthma research.

• Standardization and Reproducibility: Recombinant proteins provide batch-to-batch consistency, defined post-translational modifications, and species specificity, which are critical for reproducible and interpretable results in both basic and translational research.

Typical applications include:

• ELISA (as a capture antigen or standard)
• Flow cytometry and immunophenotyping
• Functional cell-based assays (e.g., IgE binding, B cell activation)
• Blocking and competition assays
• Structural and biophysical studies

In summary, Recombinant Human CD23 is a versatile and essential reagent for immunology, allergy, oncology, and therapeutic antibody research, enabling precise and reproducible investigation of CD23’s roles in health and disease.

Yes, recombinant human CD23 can be used as a standard for quantification or calibration in ELISA assays, provided it matches the form of CD23 you intend to measure and is compatible with your assay's antibodies and detection system.

Essential context and supporting details:

• Recombinant CD23 as Standard: Many commercial ELISA kits for human CD23 use recombinant human CD23 as their standard for generating calibration curves. This allows for accurate quantification of CD23 in biological samples by comparing sample signals to those from known concentrations of the recombinant protein.

• Form and Compatibility: Ensure the recombinant CD23 you use as a standard is of the same isoform and domain (e.g., extracellular domain, soluble CD23) as the target in your samples. Most kits use the extracellular domain or soluble form for calibration. The antibodies in your ELISA must recognize the recombinant standard equivalently to the native protein in your samples.

• Validation: The recombinant standard should be validated for use in your specific ELISA system. This includes confirming linearity, sensitivity, and specificity within your assay's dynamic range. The standard curve should be prepared freshly and run in parallel with your samples for each assay.

• Preparation and Handling: Follow best practices for reconstitution, dilution, and storage of the recombinant standard to maintain its integrity and reproducibility. Avoid mixing standards from different lots or sources unless equivalence has been established.

• Limitations: If your recombinant CD23 differs structurally from the native protein in your samples (e.g., glycosylation, isoform), this may affect quantification accuracy. Always verify that your ELISA antibodies do not discriminate between recombinant and native forms.

Summary of best practices:

• Use recombinant human CD23 that matches the form present in your samples.
• Validate the standard curve for accuracy and reproducibility.
• Prepare standards according to recommended protocols.
• Confirm antibody cross-reactivity and specificity for both recombinant and native CD23.

If you follow these guidelines, recombinant human CD23 is appropriate for use as a standard in ELISA quantification and calibration.

Recombinant Human CD23 has been validated for several applications in published research, primarily in the context of immunology, allergy, and B-cell biology. Key applications include:

• ELISA (Enzyme-Linked Immunosorbent Assay): Used as a capture protein in ELISA assays to detect IgE and other ligands, and as a standard for quantification (e.g., in allergy biomarker studies and immunotherapy monitoring).
• Blocking Experiments: Employed as a control fragment to block antibody binding in immunoassays, confirming antibody specificity in Western blot (WB), immunohistochemistry (IHC), and immunocytochemistry (ICC).
• Cell-Based Assays: Utilized in studies of B-cell signaling, IgE regulation, and receptor-ligand interactions, including assays to measure IgE binding and inhibition.
• Antibody Screening: Applied in the development and validation of anti-CD23 and anti-IgE antibodies, including therapeutic candidates.
• Bioassays: Used in functional assays to study the effects of CD23 on IgE production, B-cell differentiation, and immune modulation.
• CAR (Chimeric Antigen Receptor) Detection: Employed as a target protein in assays for CAR-T cell development and validation, particularly in models of B-cell malignancies.
• Structural and Interaction Studies: Used in studies to elucidate the structure of CD23 and its interactions with IgE, CD21, and other binding partners.

These applications are supported by research in areas such as allergic responses, B-cell malignancies, and immunotherapy, highlighting the versatility of recombinant human CD23 in both basic and translational research.

To reconstitute and prepare Recombinant Human CD23 protein for cell culture experiments, first centrifuge the vial briefly before opening, then add sterile distilled water or sterile PBS to achieve a final concentration of 0.1–0.5 mg/mL (or as specified in your experimental protocol). Avoid vigorous mixing—do not vortex or pipette forcefully, as this can denature the protein.

Step-by-step protocol:

• Centrifuge the vial briefly to collect the lyophilized powder at the bottom before opening.
• Add sterile distilled water or PBS to the vial to reach your desired concentration (commonly 0.1–0.5 mg/mL for cell culture applications).
• Gently mix by swirling or inverting the vial. Allow the protein to dissolve at room temperature for 15–30 minutes. Avoid foaming and do not vortex.
• If your application requires, add a carrier protein or stabilizer (e.g., 0.1% BSA, 5% HSA, 10% FBS, or 5% trehalose) to the reconstituted solution to enhance stability, especially for long-term storage or low concentration use.
• Aliquot the solution to minimize freeze-thaw cycles. Store aliquots at −20°C for up to 3 months, or at 2–8°C for up to 1 week.
• For cell culture, dilute the reconstituted protein into your culture medium to the desired working concentration immediately before use.

Additional notes:

• For serum-free or animal experiments, avoid using human or animal proteins as carriers; use trehalose instead.
• Always wear appropriate PPE (lab coat, gloves) when handling recombinant proteins.
• Avoid repeated freeze-thaw cycles, as these can degrade the protein.
• If the protein is to be used in sensitive assays, ensure endotoxin levels are suitable for your application (typically <0.1 EU/μg for cell culture).

Summary Table: Recombinant Human CD23 Reconstitution

This protocol ensures optimal solubility and stability of recombinant CD23 for cell culture experiments, minimizing protein loss and maintaining bioactivity.