Recombinant Human Desmoglein-1

Recombinant Human Desmoglein-1 (DSG1) is a valuable tool for research applications focused on cell-cell adhesion, epithelial biology, autoimmune disease mechanisms, and diagnostic assay development. Its use enables precise, reproducible studies of desmosomal function and related pathologies.

Key scientific reasons to use recombinant DSG1 include:

• Modeling Cell Adhesion and Epithelial Integrity: DSG1 is a critical component of desmosomes, mediating strong intercellular adhesion in epithelial tissues. Recombinant DSG1 allows for in vitro studies of desmosome assembly, cell-cell adhesion, and the molecular interactions between desmogleins, desmocollins, and cytoskeletal proteins.

• Investigating Disease Mechanisms: DSG1 dysfunction is implicated in skin diseases such as pemphigus foliaceus, where autoantibodies target DSG1, leading to loss of keratinocyte adhesion and blister formation. Recombinant DSG1 can be used to study autoantibody binding, pathogenic mechanisms, and to develop neutralization or blocking assays.

• Diagnostic Assay Development: Recombinant DSG1 is widely used as an antigen in ELISA assays for the serodiagnosis of pemphigus. These assays are sensitive and specific for detecting anti-DSG1 autoantibodies in patient sera, aiding in diagnosis and monitoring disease activity.

• Functional Studies in Signal Transduction and Differentiation: DSG1 regulates epithelial differentiation and signaling pathways, such as the extracellular signal-regulated kinase (ERK) pathway. Recombinant DSG1 enables mechanistic studies of these processes, including gene expression profiling and pathway analysis in epithelial cells.

• Control and Standardization: Recombinant DSG1 provides a consistent, well-characterized protein for use as a control in immunoassays, protein arrays, and blocking/neutralizing experiments, ensuring reproducibility and reliability in research protocols.

Typical applications include:

• ELISA and immunoassays for autoantibody detection.
• Cell culture studies to assess adhesion and signaling.
• Protein-protein interaction assays and structural studies.
• Blocking/neutralization experiments to dissect functional roles.

In summary, recombinant human DSG1 is essential for dissecting the molecular basis of epithelial adhesion, autoimmune skin diseases, and related signaling pathways, providing a robust platform for both basic and translational research.

You can use recombinant human Desmoglein-1 as a standard for quantification or calibration in your ELISA assays, provided it is of high purity and its concentration is accurately determined. This is a common and accepted practice in quantitative ELISA protocols.

Key considerations and best practices:

• Purity and Identity: The recombinant protein should be highly purified and well-characterized to ensure it represents the native antigen detected by your assay.
• Concentration Determination: Accurately determine the concentration of your recombinant Desmoglein-1 standard, ideally using a reliable method such as absorbance at 280 nm with a known extinction coefficient, or HPLC quantification.
• Standard Curve Preparation: Prepare a dilution series of the recombinant protein in the same buffer as your samples to generate a standard curve. This allows you to interpolate the concentration of Desmoglein-1 in your unknown samples based on their optical density (O.D.) values.
• Range and Sensitivity: Ensure your standard curve covers the expected concentration range of your samples and falls within the assay’s detection limits (e.g., typical ELISA kits for Desmoglein-1 have standard curves ranging from ~0.3–20 ng/mL or up to 200 ng/mL, depending on the kit).
• Matrix Effects: If your samples are in a complex matrix (e.g., serum, plasma), consider preparing your standards in the same matrix or using appropriate diluents to minimize matrix effects and improve quantification accuracy.
• Validation: If you are using a recombinant standard not provided with a commercial kit, validate its performance in your specific ELISA system. This includes confirming parallelism between the standard curve and sample dilution curves, and ensuring the recombinant protein is recognized equivalently by the assay antibodies.

Protocol references:

• Most ELISA kits and protocols recommend reconstituting the standard (often recombinant protein) and preparing a serial dilution to generate a standard curve for quantification.
• The use of recombinant proteins as standards is standard practice when purified native protein is unavailable or impractical to obtain.

Summary:
Using recombinant human Desmoglein-1 as a standard is scientifically valid for ELISA quantification, as long as you ensure its purity, accurate concentration, and compatibility with your assay system. Always validate the standard in your specific assay context for reliable results.

Recombinant Human Desmoglein-1 has been validated for several key applications in published research, most notably in ELISA-based serodiagnosis of pemphigus, studies of cell adhesion and epithelial barrier function, and as a control or blocking reagent in immunological assays.

Validated Applications in Published Research:

• ELISA (Enzyme-Linked Immunosorbent Assay):

  • Recombinant Desmoglein-1 is widely used as an antigen in ELISA systems for the serodiagnosis of autoimmune blistering diseases, especially pemphigus foliaceus and pemphigus vulgaris. These assays detect circulating anti-Desmoglein-1 autoantibodies in patient sera and are considered sensitive and specific for clinical diagnosis and disease monitoring.
  • The recombinant protein expressed in insect cells has been shown to outperform indirect immunofluorescence microscopy for antibody detection.

• Cell Adhesion and Barrier Function Studies:

  • Recombinant Desmoglein-1 is used in research to study its role in epithelial cell-cell adhesion, desmosome formation, and maintenance of tissue integrity. For example, it has been employed in in vitro models to investigate the impact of Desmoglein-1 dysregulation on esophageal epithelial barrier function and its involvement in diseases such as eosinophilic esophagitis and atopic dermatitis.
  • Functional studies often utilize recombinant protein to probe signaling pathways and cellular responses related to desmosomal cadherins.

• Blocking/Neutralizing and Control Experiments:

  • The protein is used as a blocking or neutralizing agent in immunological assays to confirm antibody specificity or to inhibit Desmoglein-1-mediated cell adhesion.
  • It serves as a control protein in various experimental setups, including protein arrays and immunoassays.

• Protein Array and Signal Transduction Research:

  • Recombinant Desmoglein-1 is included in protein arrays for profiling autoantibody responses and investigating signal transduction pathways involving desmosomal proteins.

Additional Notes:

• Recombinant Desmoglein-1 is not intended for therapeutic or diagnostic use in humans but is validated for research purposes.
• It is commonly used in dermatological research, particularly in studies of autoimmune skin diseases and epithelial biology.

Summary Table of Validated Applications

If you require protocols or technical details for any specific application, please specify the intended use.

To reconstitute and prepare Recombinant Human Desmoglein-1 protein for cell culture experiments, first briefly centrifuge the vial to collect the lyophilized powder at the bottom, then add sterile deionized water or buffer to achieve a concentration of 0.1–1.0 mg/mL, gently mix, and avoid vigorous agitation. For optimal stability and activity, consider adding 5–50% glycerol and aliquot for storage at –20°C or –80°C.

Detailed protocol:

• Equilibrate the vial and reconstitution buffer to room temperature before opening.
• Centrifuge the vial briefly to ensure all powder is at the bottom.
• Add reconstitution buffer:
  • Most protocols recommend sterile deionized water for initial reconstitution.
  • Some preparations may allow or recommend sterile PBS or a specific buffer (e.g., 20 mM Tris, 150 mM NaCl, pH 8.0) if higher stability or compatibility with downstream applications is needed. Always check the product datasheet for buffer compatibility.
• Target concentration: 0.1–1.0 mg/mL is standard for storage; adjust as needed for your application.
• Mix gently by pipetting up and down or gentle swirling. Do not vortex or shake vigorously, as this can denature the protein.
• Allow to dissolve at room temperature for 15–30 minutes. The solution should become clear.
• Aliquot the reconstituted protein into small volumes to avoid repeated freeze-thaw cycles.
• Additives for stability: For long-term storage, add 5–50% glycerol (final concentration) to prevent freeze-thaw damage.
• Storage: Store aliquots at –20°C or –80°C. For short-term use (up to 1 month), 2–8°C is acceptable.

For cell culture use:

• Before adding to cells, dilute the protein to the desired working concentration using cell culture medium or buffer containing carrier proteins (e.g., 0.1% BSA, 5% HSA, or 10% FBS) to minimize adsorption and stabilize the protein.
• Do not dilute directly in water for cell culture applications, as this may cause protein degradation.

Summary of key points:

• Use sterile technique throughout.
• Reconstitute at 0.1–1.0 mg/mL in sterile water or buffer.
• Add glycerol for storage stability.
• Aliquot and store at –20°C or –80°C.
• Dilute to working concentration in cell culture medium with carrier protein before use.

Always consult the specific product datasheet or Certificate of Analysis for any unique requirements for your recombinant Desmoglein-1 preparation.