Recombinant Mouse EDAR

Recombinant Mouse EDAR is used in research applications to study the molecular mechanisms of ectodermal development, disease modeling, and signaling pathways relevant to both mouse and human biology. Its recombinant form allows for controlled, reproducible experiments and enables detailed functional analyses of EDAR-mediated processes.

Key scientific reasons to use recombinant Mouse EDAR include:

• Modeling Human Disease and Evolution: Mouse EDAR is structurally and functionally similar to human EDAR, making it a valuable tool for modeling human genetic variants and their phenotypic consequences, such as hair thickness, gland development, and mammary gland structure. Recombinant EDAR enables precise manipulation and study of these traits in vitro and in vivo.

• Tumor Suppressor Studies: EDAR acts as a tumor suppressor in melanoma by inducing apoptosis in the absence of its ligand. Recombinant EDAR can be used to investigate its role in cancer biology, including its pro-apoptotic activity and impact on tumor progression.

• Ectodermal Development Research: EDAR signaling is essential for the normal development of ectodermal derivatives, including hair follicles, teeth, and glands. Recombinant EDAR allows for the dissection of signaling pathways and the identification of downstream effectors involved in these developmental processes.

• Controlled and Reproducible Experiments: Recombinant proteins offer high purity, batch-to-batch consistency, and defined activity, which are critical for quantitative assays, mechanistic studies, and therapeutic screening. This ensures reliable data and facilitates comparison across experiments.

• Pharmacological and Functional Assays: Recombinant EDAR can be used to stimulate or inhibit signaling pathways in cell-based assays, enabling the study of downstream effects such as NFκB activation, apoptosis, or differentiation.

• Translational Research: Findings from mouse EDAR studies can inform human association studies and therapeutic development, given the high degree of conservation between mouse and human EDAR death domains and signaling mechanisms.

In summary, recombinant Mouse EDAR is a versatile tool for investigating developmental biology, disease mechanisms, and therapeutic strategies, offering precise control and relevance to both basic and translational research.

You can use recombinant Mouse EDAR as a standard for quantification or calibration in your ELISA assays, provided that the recombinant protein is of high purity, its concentration is accurately determined, and it is compatible with your assay format.

Key considerations and supporting details:

• Purity and Quantification: The recombinant protein used as a standard should be highly purified, and its concentration must be precisely measured, ideally by methods such as HPLC or absorbance at 280 nm. Impurities or inaccurate quantification can lead to unreliable standard curves and assay results.

• Standard Curve Preparation: Prepare a serial dilution of the recombinant Mouse EDAR to generate a standard curve in each assay. This allows for the quantification of EDAR in your samples by comparison to the known concentrations of the standard.

• Matrix Effects: If your samples are in a complex matrix (e.g., serum, plasma), perform spike-and-recovery and linearity experiments to ensure that the recombinant standard behaves similarly to endogenous EDAR in your assay conditions. This helps validate that the standard curve accurately reflects the analyte in your sample type.

• Reference Standards: For best practice and harmonization, use standards calibrated to international reference materials (such as NIBSC or WHO standards) if available. If such reference standards are not available for Mouse EDAR, your recombinant protein can serve as a practical standard, but results may not be directly comparable across different laboratories or assay platforms.

• Assay Validation: Validate the use of recombinant Mouse EDAR as a standard in your specific ELISA system by assessing accuracy, precision, and reproducibility across multiple runs and sample types.

• ELISA Kit Compatibility: Some commercial ELISA kits are designed to detect native, not recombinant, EDAR, and may not be validated for use with recombinant standards. Check the specificity and compatibility of your antibodies and assay format with recombinant EDAR.

• Controls: Include appropriate controls, such as endogenous positive controls, to ensure assay reliability when using recombinant proteins as standards.

In summary, recombinant Mouse EDAR is suitable as a standard for ELISA quantification if it is well-characterized, pure, and validated for your assay conditions. Always confirm compatibility with your specific assay reagents and sample types, and perform necessary validation experiments to ensure accurate quantification.

Recombinant Mouse EDAR has been validated in published research primarily for applications involving the study of ectodermal development, signaling pathway analysis, and functional rescue experiments in both in vitro and in vivo models.

Key validated applications include:

• In vivo functional studies: Recombinant mouse EDAR has been used to generate knock-in mouse models to study the phenotypic consequences of specific EDAR variants (such as EDARV370A), including effects on hair thickness, eccrine gland number, mammary gland structure, and other ectodermal traits. These studies directly assess the biological function of EDAR in a physiological context.

• In vitro signaling assays: Overexpression of recombinant EDAR (including variant forms) in cell culture has been used to analyze downstream signaling, particularly NFκB pathway activation, to compare the functional impact of different EDAR alleles.

• Rescue experiments: Recombinant EDAR has been applied in genetic rescue experiments, such as introducing gain-of-function EDAR alleles into loss-of-function backgrounds to determine the sufficiency of EDAR signaling for restoring normal development of skin appendages and glands.

• Morphological and functional correction studies: Pharmacological stimulation of EDAR signaling, sometimes using recombinant EDAR agonists or antibodies, has been validated for correcting morphological and functional defects in adult mouse tissues, such as sebaceous glands, demonstrating the utility of recombinant EDAR in therapeutic modeling.

• Ex vivo organ culture and patterning assays: Recombinant EDAR has been used in reconstituted skin and organ culture systems to study its role in the patterning and development of ectodermal appendages, such as feather buds and hair follicles.

• Protein detection and quantification: Recombinant mouse EDAR is used as a standard or control in direct ELISA and Western blot assays to detect and quantify EDAR protein levels, and to assess antibody specificity.

These applications demonstrate that recombinant mouse EDAR is a versatile tool for dissecting the molecular and developmental roles of EDAR signaling in mammalian biology, with validation in both basic research and translational contexts.

To reconstitute and prepare Recombinant Mouse EDAR protein for cell culture experiments, dissolve the lyophilized protein in sterile buffer (typically PBS, pH 7.4) to a concentration of at least 100 μg/mL, ideally with a carrier protein such as 0.1% BSA to enhance stability 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 buffer: Use sterile PBS (pH 7.4) as the reconstitution buffer. If the protein was lyophilized with stabilizers (e.g., trehalose, mannitol, Tween 80), PBS alone is sufficient. If not, or for added stability, supplement with 0.1% BSA.
• Reconstitution concentration: Do not reconstitute below 100 μg/mL to minimize loss from adsorption and to maintain protein stability.
• Dissolve gently: Allow the protein to dissolve at room temperature for 15–30 minutes with gentle agitation. Avoid vigorous shaking or vortexing to prevent foaming and denaturation.
• Aliquot and storage: After reconstitution, aliquot the solution to avoid repeated freeze-thaw cycles. Store aliquots at 4–8 °C for short-term use (2–7 days) or at –20 °C to –80 °C for long-term storage (up to 3 months).
• Sterility: Always use sterile technique and sterile buffers to prevent contamination, especially for cell culture applications.

For cell culture experiments:

• Dilute the reconstituted stock to the desired working concentration using sterile culture medium immediately before use.
• Carrier protein: If your application is sensitive to serum or BSA, use a low-protein binding tube and consider adding a non-interfering carrier such as 0.1% BSA or 0.01% Tween 80, as present in the lyophilized formulation.
• Avoid repeated freeze-thaw cycles to maintain protein integrity.

Summary of best practices:

• Use sterile PBS (pH 7.4) or PBS + 0.1% BSA for reconstitution.
• Reconstitute to ≥100 μg/mL.
• Dissolve gently at room temperature.
• Aliquot and store at –20 °C or below for long-term use.
• Dilute into cell culture medium just before use.

If your specific product includes a printed manual, always follow those instructions for optimal results.