Recombinant Human VEGF₁₂₁ (aa 207-318)

Recombinant Human VEGF₁₂₁ (aa 207-318) is widely used in research because it is a potent, well-characterized isoform of vascular endothelial growth factor (VEGF) that robustly stimulates angiogenesis, endothelial cell proliferation, migration, and survival, making it ideal for studies of vascular biology, tissue regeneration, and related therapeutic applications.

Key reasons to use this protein in your research:

• Potent Angiogenic Activity: VEGF₁₂₁ strongly promotes the proliferation, migration, and tube formation of endothelial cells, which are critical steps in angiogenesis. In vitro studies show significant enhancement of human umbilical vein endothelial cell (HUVEC) proliferation and wound healing when treated with recombinant VEGF₁₂₁.
• Defined Structure and Bioactivity: The 121-amino acid isoform (aa 207-318 of the precursor) is the shortest major VEGF-A splice variant, lacking heparin-binding domains, which makes it freely diffusible and suitable for studies where uniform distribution is desired. It retains full biological activity, binding to VEGF receptors (VEGFR1/Flt-1 and VEGFR2/KDR) and co-receptors such as neuropilin-1, thereby activating downstream signaling pathways.
• Reproducibility and Consistency: Recombinant production ensures batch-to-batch consistency, purity, and defined activity, which are essential for quantitative and mechanistic studies.
• Broad Application Range: VEGF₁₂₁ is used in:
  • Angiogenesis assays (e.g., tube formation, migration, and proliferation assays in endothelial cells).
  • Wound healing models (accelerates closure and vascularization).
  • Neurobiology (supports survival and growth of neurons and astrocytes).
  • Tissue engineering and regenerative medicine (promotes vascularization of engineered tissues).
  • Disease modeling (studying pathological angiogenesis in cancer, retinopathies, and preeclampsia).
• Cross-Species Reactivity: Human VEGF₁₂₁ shares high sequence identity with VEGF from other mammals, making it suitable for use in various animal models.

Summary of scientific findings:

• VEGF₁₂₁ stimulates HUVEC proliferation in a dose-dependent manner (ED₅₀ ≈ 0.75–3.75 ng/mL).
• It accelerates wound healing and enhances blood vessel formation in vitro and in vivo models.
• It is biologically active in multiple assay systems, including bioassays, binding assays, and surface plasmon resonance.

In conclusion, recombinant human VEGF₁₂₁ (aa 207-318) is a gold-standard reagent for angiogenesis and vascular biology research due to its robust, reproducible activity and broad utility in both basic and translational studies.

Yes, recombinant human VEGF₁₂₁ (aa 207-318) can be used as a standard for quantification or calibration in ELISA assays, provided it matches the isoform and epitope recognized by your assay antibodies. This isoform is commonly used as an ELISA standard, but there are important considerations to ensure accuracy and compatibility.

Key considerations and supporting details:

• Isoform and Sequence Compatibility: VEGF₁₂₁ (aa 207-318) corresponds to the mature, secreted form of human VEGF₁₂₁, which is widely used as a standard in ELISA assays targeting VEGF-A. However, some commercial ELISA kits use VEGF₁₆₅ as the calibrator, which has a longer sequence (Ala27–Arg191) and includes additional exons not present in VEGF₁₂₁. Ensure your assay is validated for VEGF₁₂₁ or that the antibodies used recognize epitopes present in the 121 isoform.

• Carrier Protein: Recombinant VEGF₁₂₁ is available in both carrier-free and BSA-containing formulations. For ELISA standards, the BSA-containing form is generally recommended to enhance stability and reproducibility. Carrier-free protein is preferred only if BSA interferes with your assay.

• Dimerization and Activity: VEGF₁₂₁ functions as a disulfide-linked homodimer, and recombinant preparations should maintain this structure for accurate quantification. Confirm that your recombinant protein is properly folded and dimerized, as monomeric or misfolded protein may not be recognized equivalently by ELISA antibodies.

• Validation and Parallelism: When using a recombinant standard, it is essential to verify that the standard curve generated is parallel to the response of endogenous VEGF in your samples. This ensures that the recombinant standard behaves similarly to native VEGF in your assay matrix.

• Documentation and Protocols: Always consult your ELISA kit’s documentation to confirm which VEGF isoform and sequence are recommended as standards. Some kits specify the use of VEGF₁₆₅, while others are validated for VEGF₁₂₁.

Summary Table: VEGF ELISA Standard Considerations

Best Practices:

• Confirm that your ELISA antibodies detect VEGF₁₂₁ (aa 207-318).
• Use the same isoform as the kit standard for calibration whenever possible.
• Validate parallelism between your recombinant standard and native VEGF in your sample matrix.
• Use BSA-containing formulations unless BSA interferes with your assay.

If your ELISA kit or protocol specifically calls for VEGF₁₂₁ (aa 207-318), this recombinant protein is appropriate as a standard. If the kit uses a different isoform (e.g., VEGF₁₆₅), results may not be directly comparable unless validated for cross-isoform quantification.

Recombinant Human VEGF₁₂₁ (aa 207-318) has been validated for several key applications in published research, primarily related to angiogenesis and endothelial cell biology.

Validated Applications:

• Bioassays for Angiogenic Activity:
  VEGF₁₂₁ (aa 207-318) stimulates proliferation, migration, and tube formation in human umbilical vein endothelial cells (HUVECs). These assays are standard for evaluating angiogenic potential and endothelial cell function.

• Wound Healing (Migration) Assays:
  Recombinant VEGF₁₂₁ has been used in in vitro wound healing (scratch) assays to assess its ability to promote endothelial cell migration and wound closure.

• Tube Formation Assays:
  The protein induces capillary-like tube formation in HUVECs, a hallmark of angiogenic activity.

• Chorioallantoic Membrane (CAM) Assay:
  Proangiogenic effects have been validated ex ovo using the chicken CAM assay, demonstrating its ability to stimulate new blood vessel formation in embryonated eggs.

• Cell Proliferation Assays:
  Dose-dependent stimulation of HUVEC proliferation has been reported, with ED₅₀ values in the low ng/mL range.

• Binding Assays:
  VEGF₁₂₁ has been used in binding assays to study interactions with receptors such as VEGFR1, VEGFR2, and neuropilin-1.

• ELISA Standard:
  The recombinant protein is validated as a standard in ELISA for quantifying VEGF levels.

Additional Context:

• Surface Plasmon Resonance (SPR):
  Used to characterize receptor binding kinetics.

• Matrix Preparation:
  VEGF₁₂₁ has been incorporated into cellular matrices for tissue engineering applications.

• Therapeutic Delivery Studies:
  Encapsulation in poly-l-lactide (PLA) microparticles for sustained release and therapeutic angiogenesis has been validated.

Summary Table of Applications

Key Insights:

• VEGF₁₂₁ (aa 207-318) is widely used for in vitro and ex ovo angiogenesis studies, wound healing, and as a standard for VEGF quantification.
• Its activity is validated in both direct cell-based assays and in delivery systems for therapeutic research.
• The protein is suitable for mechanistic studies involving VEGF receptor interactions and downstream signaling.

If you require protocols or specific experimental details for any of these applications, please specify the assay or context.

To reconstitute and prepare Recombinant Human VEGF₁₂₁ (aa 207-318) for cell culture experiments, dissolve the lyophilized protein in sterile phosphate-buffered saline (PBS) containing at least 0.1% human or bovine serum albumin (BSA or HSA) to a final concentration of 50 µg/mL or higher. This carrier protein is essential to stabilize VEGF and prevent adsorption to surfaces.

Step-by-step protocol:

• Centrifuge the vial briefly before opening to collect all lyophilized material at the bottom.
• Add sterile PBS with 0.1% BSA/HSA to the vial to reach the desired concentration (≥50 µg/mL).
• Gently pipette the solution up and down along the vial wall to dissolve the protein completely. Do not vortex.
• Allow the solution to sit for a few minutes at room temperature to ensure full dissolution.
• If needed, further dilute the reconstituted VEGF₁₂₁ in cell culture medium or PBS with 0.1% BSA/HSA to reach your working concentration (typical bioactive range: 0.75–3.75 ng/mL for HUVEC proliferation assays).

Storage and handling:

• Aliquot the reconstituted protein to avoid repeated freeze-thaw cycles.
• Store aliquots at –20 °C to –80 °C for long-term storage; at 2–8 °C for up to 3 months for short-term use.
• Avoid multiple freeze-thaw cycles, as this can reduce protein activity.

Additional notes:

• If your protocol or supplier recommends, you may use sterile water for initial reconstitution, but PBS with carrier protein is preferred for stability and compatibility with cell culture.
• Always consult the product-specific datasheet for any unique instructions, as formulations may vary.

Summary of key points:

• Reconstitute at ≥50 µg/mL in sterile PBS + 0.1% BSA/HSA
• Gently pipette to dissolve, do not vortex
• Aliquot and store at –20 °C or below
• Avoid repeated freeze-thaw cycles

This protocol ensures optimal stability and bioactivity of VEGF₁₂₁ for cell culture applications.