Recombinant Human VEGF-B

Recombinant Human VEGF-B is used in research applications primarily for its roles in cell survival, anti-angiogenesis, metabolic regulation, and tissue protection, making it a valuable tool for studies in cardiovascular biology, neuroprotection, cancer, and metabolic disease.

Key scientific applications and rationale include:

• Cell Survival and Anti-Apoptosis: VEGF-B is a potent survival factor for various cell types, including neurons and cardiomyocytes. It inhibits apoptosis by suppressing pro-apoptotic proteins, making it useful for neuroprotection and studies of cell death mechanisms. For example, recombinant VEGF-B protects retinal and motor neurons from apoptosis and can prolong survival in neurodegeneration models.

• Anti-Angiogenic Activity: Unlike other VEGF family members, VEGF-B can inhibit excessive angiogenesis, particularly by antagonizing the FGF2/FGFR1 pathway. This property is relevant for cancer research, where VEGF-B has been shown to restrict tumor angiogenesis and growth in various models. High VEGF-B levels correlate with reduced tumor angiogenesis and better survival outcomes in humans.

• Cardiovascular and Metabolic Effects: Recombinant VEGF-B increases coronary vasculature, improves tissue perfusion, and reprograms metabolic pathways in the heart and adipose tissue. It has therapeutic potential for protecting the heart from ischemic injury, improving metabolic health, and preventing obesity-related complications. VEGF-B signaling enhances capillary density and insulin supply in adipose tissue, supporting studies in diabetes and metabolic syndrome.

• Fibrosis and Tissue Remodeling: VEGF-B attenuates activation of cardiac fibroblasts and reduces fibrosis in aging hearts, suggesting applications in cardiac remodeling and fibrosis research.

• Non-Angiogenic Therapeutic Potential: Fusion proteins and modified forms of VEGF-B (e.g., ELP-VEGF-B) are being explored for conditions like preeclampsia, where non-angiogenic, tissue-protective effects are desirable.

Best practices for use:

• Select the appropriate isoform and formulation (e.g., carrier-free for biochemical assays, BSA-containing for cell culture) based on your experimental needs.
• Validate biological activity in relevant cell types (e.g., endothelial cells, neurons, fibroblasts) using functional assays such as cell survival, migration, or angiogenesis assays.

Summary of research applications:

• Neuroprotection and anti-apoptosis studies
• Cancer and tumor angiogenesis inhibition
• Cardiovascular protection and metabolic regulation
• Fibrosis and tissue remodeling models
• Investigating VEGF-B signaling pathways

Using recombinant human VEGF-B enables precise mechanistic studies and therapeutic modeling in these areas, supporting both basic research and translational applications.

Recombinant Human VEGF-B can be used as a standard for quantification or calibration in ELISA assays, provided the assay is specifically designed to detect VEGF-B and the recombinant protein matches the isoform and sequence recognized by the assay antibodies.

For accurate quantification:

• Ensure your ELISA kit or custom assay is validated for both natural and recombinant human VEGF-B. Most commercial VEGF-B ELISA kits are designed to recognize both forms, allowing recombinant VEGF-B to serve as a standard for calibration.
• The recombinant protein should be of high purity and formulated appropriately (often with BSA for stability), as recommended for use as an ELISA standard.
• Prepare a standard curve using serial dilutions of the recombinant VEGF-B within the dynamic range specified by your assay protocol (e.g., 15.63–1000 pg/mL for some kits).
• Confirm that the recombinant VEGF-B isoform (e.g., VEGF-B167 or VEGF-B186) matches the target of your assay antibodies, as cross-reactivity with other VEGF family members is generally low.

Important technical notes:

• Do not use recombinant VEGF-A or VEGF165 as a standard for VEGF-B quantification, as ELISA kits for VEGF-A do not recognize VEGF-B due to low sequence homology.
• Always verify that your recombinant VEGF-B standard is compatible with the antibody pair used in your ELISA, as some kits are isoform-specific.
• Follow best practices for protein handling: avoid repeated freeze/thaw cycles and use freshly prepared dilutions for each assay run.

In summary, recombinant human VEGF-B is suitable as a standard for ELISA quantification of VEGF-B, provided the assay is validated for this application and the recombinant protein matches the assay’s specificity requirements.

Recombinant Human VEGF-B has been validated for several key applications in published research, primarily in the fields of neurobiology, cell biology, and functional assays:

• Functional Assays: VEGF-B is widely used in in vitro and in vivo functional assays to study its biological effects, such as promoting nerve regeneration, cell survival, and angiogenesis.
• ELISA (Enzyme-Linked Immunosorbent Assay): It is validated for use in ELISA to quantify VEGF-B protein levels in biological samples.
• Western Blot: VEGF-B has been used in Western blotting to detect its expression and analyze its molecular weight and purity.

Published Research Applications

• Peripheral Nerve Regeneration: VEGF-B has been shown to stimulate regeneration of injured peripheral neurons, enhance sensory recovery, and promote tissue healing in animal models. These effects are mediated through PI3K and Notch signaling pathways.
• Neuroprotection: VEGF-B treatment rescues neurons from apoptosis in the retina and brain without inducing undesired angiogenesis, making it a promising neuroprotective agent.
• Stroke and Ischemic Injury: Administration of VEGF-B improves recovery from cerebral ischemic injury and reduces stroke volume in animal models.
• Angiogenesis and Endothelial Cell Biology: VEGF-B acts as a growth factor for endothelial cells and has been studied for its role in angiogenesis and vascular maintenance.
• Stem Cell Research: VEGF-B is used to support endothelial cell growth and maintenance in stem cell differentiation protocols.
• Bioactivity Assays: Recombinant VEGF-B is routinely validated in bioactivity assays to confirm its functional properties, such as promoting cell survival and proliferation.

Additional Notes

• Non-Angiogenic Therapeutic Potential: Unlike VEGF-A, VEGF-B does not strongly induce angiogenesis, making it suitable for therapeutic applications where vascular proliferation is undesirable, such as in preeclampsia and neurodegenerative diseases.
• Species and Sample Types: VEGF-B has been validated in human and mouse models, and used with various sample types including whole cells and tissue spheroids.

In summary, Recombinant Human VEGF-B is validated for use in functional assays, ELISA, and Western blot, and is extensively applied in research on nerve regeneration, neuroprotection, angiogenesis, and stem cell biology.

Reconstitution of Recombinant Human VEGF-B Protein

Recombinant Human VEGF-B proteins are typically supplied as lyophilized (freeze-dried) material and require proper reconstitution before use in cell culture experiments. The reconstitution process is critical for maintaining protein stability and bioactivity.

Reconstitution Protocol

Initial Preparation

Begin by allowing the lyophilized vial to reach room temperature before opening. This prevents condensation from entering the vial. When reconstituting, use sterile, distilled water as the primary diluent, unless the product documentation specifies an alternative. For optimal results, avoid vigorous shaking or vortexing during reconstitution, as this can cause foaming and protein denaturation.

Reconstitution Procedure

Add the appropriate volume of sterile distilled water to achieve your desired concentration. A common reconstitution concentration is 500 μg/mL in sterile deionized water, though specific volumes depend on the vial size and your experimental requirements. After adding the diluent, allow the vial to reconstitute for 15-30 minutes at room temperature with gentle agitation. Gently mix the vial by swirling or inverting several times to ensure complete and homogeneous dissolution of the protein powder.

Carrier Protein Considerations

For long-term storage and to prevent protein aggregation, it is recommended to add a carrier protein such as bovine serum albumin (BSA) at a concentration of 0.1% to your reconstituted solution. This is particularly important if you plan to store aliquots for extended periods or perform multiple freeze-thaw cycles.

Storage and Stability

Short-term Storage

After reconstitution under aseptic conditions, the protein may be stored at 2–8°C for up to one month. This temperature range is suitable for experiments planned within a short timeframe.

Long-term Storage

For extended storage, maintain the reconstituted protein at −20°C to −70°C in a manual defrost freezer. Avoid repeated freeze-thaw cycles, as these can compromise protein integrity and bioactivity. If you anticipate multiple uses, prepare small aliquots to minimize handling and freeze-thaw exposure.

Stability Timeline

Reconstituted VEGF-B protein is typically guaranteed for 3 months from the date of reconstitution or until the expiration date listed on the product documentation, whichever comes first.

Practical Recommendations for Cell Culture

When preparing VEGF-B for cell culture applications, ensure all reconstitution is performed under aseptic conditions to maintain sterility. Prepare fresh standard dilutions immediately prior to use if you are conducting dose-response experiments. The reconstituted protein should be gently mixed before each use, and any unused portions should be promptly returned to appropriate storage conditions. Given that VEGF-B exists as a disulfide-linked homodimeric protein, maintaining proper storage conditions is essential to preserve the native dimeric structure required for receptor binding and bioactivity.