Recombinant Mouse VEGF R2

Using Recombinant Mouse VEGF R2 (VEGFR-2/Flk-1/KDR) in research applications is essential for studying angiogenesis, tumor biology, immune modulation, and therapeutic targeting in mouse models. VEGFR-2 is the principal receptor mediating the effects of VEGF on endothelial cell proliferation, migration, and differentiation, making it a critical tool for dissecting VEGF-driven pathways in both physiological and pathological contexts.

Key scientific applications and rationales include:

• Angiogenesis Research: VEGFR-2 is the main mediator of VEGF-induced angiogenic signaling in endothelial cells. Recombinant mouse VEGFR-2 enables in vitro and in vivo studies of angiogenesis, vessel permeability, and endothelial cell function, which are central to cancer, wound healing, and developmental biology research.

• Tumor Biology and Immunology: VEGFR-2 is overexpressed in tumor vasculature and is a target for anti-angiogenic therapies. Using recombinant VEGFR-2 allows for the evaluation of inhibitors, antibodies, or engineered immune cells (such as CAR-T cells) targeting this receptor, facilitating preclinical studies on tumor growth, metastasis, and immune cell infiltration.

• Immunotherapy Studies: Blockade or modulation of VEGFR-2 can relieve VEGF-induced immune suppression, enhance T cell infiltration, and improve the efficacy of immune checkpoint blockade therapies. Recombinant VEGFR-2 is used to characterize these mechanisms and to screen for agents that modulate the tumor immune microenvironment.

• Drug Screening and Mechanistic Studies: Recombinant VEGFR-2 serves as a target for screening small molecules, antibodies, or biologics that inhibit VEGF signaling. It is also used in binding assays, structural studies, and to generate neutralizing antibodies or receptor fragments for therapeutic development.

• Species-Specific and Cross-Reactivity Studies: Mouse VEGFR-2 is required for experiments in murine models, which are standard in preclinical research. Recombinant forms allow for precise, species-matched studies, and cross-reactive reagents can bridge mouse and human research for translational applications.

• Functional Assays: Recombinant VEGFR-2 is used in ELISA, Western blot, flow cytometry, and cell-based assays to quantify ligand binding, receptor activation, and downstream signaling events.

In summary, Recombinant Mouse VEGF R2 is a versatile and indispensable reagent for mechanistic, translational, and therapeutic research involving angiogenesis, tumor progression, and immune regulation in mouse models.

Yes, recombinant Mouse VEGF R2 protein can be used as a standard for quantification or calibration in ELISA assays, provided it is properly validated and matched to the assay system.

Supporting details:

• ELISA kits for Mouse VEGF R2 are routinely validated to quantify both natural and recombinant forms of the protein. For example, sandwich ELISA kits specifically state that they can accurately measure recombinant Mouse VEGF R2, and standard curves generated with recombinant protein yield linear, reliable quantification.
• Standard preparation: Recombinant Mouse VEGF R2 is typically reconstituted in sample diluent to a known concentration, then serially diluted to generate a standard curve. The absorbance values from these standards are used to interpolate concentrations in unknown samples.
• Linearity and parallelism: Validation data from commercial ELISA kits show that recombinant standards produce dose-response curves parallel to those obtained with natural samples, confirming suitability for calibration.
• Formulation considerations: If using recombinant protein as a standard, ensure it is in a compatible formulation (e.g., with or without carrier proteins like BSA) to avoid matrix effects that could impact assay accuracy.

Best practices:

• Use recombinant Mouse VEGF R2 that matches the extracellular domain recognized by the ELISA antibodies.
• Confirm the standard’s purity and concentration using orthogonal methods (e.g., Bradford assay, spectrophotometry) before use.
• Always prepare a fresh standard curve for each assay run and verify that sample values fall within the linear range of the curve.
• Validate that the recombinant standard behaves similarly to endogenous VEGF R2 in your sample matrix by testing dilution linearity and parallelism.

Limitations:

• Recombinant standards may differ in glycosylation or folding compared to native protein, but most ELISA kits are designed to recognize both forms equivalently.
• For regulatory or clinical applications, additional validation may be required to ensure equivalence between recombinant and native standards.

In summary, recombinant Mouse VEGF R2 is appropriate for use as an ELISA standard for quantification and calibration, provided it is validated for your specific assay and sample type.

Recombinant Mouse VEGF R2 (VEGFR2/Flk-1/KDR) has been validated in published research for several key applications, primarily in the study of angiogenesis, tumor biology, and vascular signaling.

Key validated applications include:

• Bioassays: Used to measure its ability to inhibit VEGF-dependent proliferation of human umbilical vein endothelial cells (HUVECs), with reported ED50 values in the range of 10–30 ng/mL.
• Binding Assays: Employed in competitive binding assays to assess the interaction between VEGF ligands and VEGFR2, as well as to evaluate the efficacy of antibodies or inhibitors targeting VEGFR2.
• Western Blot: Utilized for detection and quantification of VEGFR2 protein expression in tissue or cell lysates, particularly in studies of tumor models and anti-angiogenic therapies.
• In Vivo Functional Studies: Applied in mouse models to investigate the role of VEGFR2 in tumor growth, angiogenesis, and metastasis, as well as to evaluate the efficacy of anti-VEGFR2 therapies (e.g., monoclonal antibodies, CAR-T cells).
• Surface Plasmon Resonance (SPR): Used to characterize binding kinetics and affinities of VEGFR2 with various ligands or therapeutic candidates.
• Gene Therapy and Immunotherapy Research: Recombinant VEGFR2 has been used as a target antigen for the development and validation of chimeric antigen receptor (CAR) T cells in mouse models of cancer.

Additional context:

• Recombinant mouse VEGFR2 is a critical tool for dissecting VEGF signaling pathways in both physiological (e.g., cardiac hypertrophy, vascular development) and pathological (e.g., tumor angiogenesis, ocular neovascularization) contexts.
• It is also used in the development and validation of novel anti-angiogenic agents, including monoclonal antibodies and bispecific antibodies, in both in vitro and in vivo settings.

Summary of main validated applications:

These applications are well-supported in the literature and form the basis for ongoing research into VEGF/VEGFR2 signaling and therapeutic targeting.

To reconstitute and prepare Recombinant Mouse VEGF R2 (VEGFR2/Flk-1) protein for cell culture experiments, dissolve the lyophilized protein in sterile PBS containing at least 0.1% serum albumin (human or bovine) to a concentration of 50–100 μg/mL. This helps maintain protein stability and activity.

Step-by-step protocol:

• Before opening the vial: Briefly centrifuge to collect all lyophilized material at the bottom.
• Reconstitution:
  • Add sterile PBS (pH 7.2–7.4) containing at least 0.1% BSA (bovine serum albumin) or HSA (human serum albumin) to achieve a final concentration of 50–100 μg/mL.
  • Gently mix by pipetting up and down or inverting the tube. Avoid vigorous vortexing, which may denature the protein.
  • Allow the solution to sit at room temperature for 10 minutes to ensure complete dissolution.
• Aliquoting and storage:
  • Divide the reconstituted solution into small aliquots to avoid repeated freeze-thaw cycles.
  • Store aliquots at ≤ −20°C for long-term use. Avoid frost-free freezers, which can cause temperature fluctuations.
• Working dilutions:
  • For cell culture, further dilute the stock solution in cell culture medium containing FBS or tissue culture grade BSA to minimize adsorption and maintain bioactivity.
  • The optimal working concentration should be determined by dose-response assays specific to your experimental system.

Additional notes:

• Always consult the product’s Certificate of Analysis (CoA) or datasheet for any manufacturer-specific instructions regarding buffer composition, concentration, and storage.
• If the protein is carrier-free, adding BSA during reconstitution is especially important to prevent loss due to adsorption to plasticware.
• Minimize freeze-thaw cycles to preserve protein integrity.

Summary Table:

This protocol ensures the recombinant VEGFR2 protein remains stable and active for cell culture applications.