Recombinant Rat FGF-9

Recombinant Rat FGF-9 is widely used in research due to its critical roles in regulating embryonic development, cell proliferation, differentiation, migration, and tissue-specific survival. Its recombinant form provides a consistent, bioactive protein source for controlled experimental applications.

Key scientific reasons to use recombinant rat FGF-9 include:

• Neurobiology: FGF-9 acts as a neurotrophic factor, supporting the growth and survival of motor neurons and glial cells, making it valuable for studies on neural development, neuroprotection, and neuroregeneration.
• Developmental Biology: It is essential for embryonic development, particularly in the regulation of cell fate decisions, organogenesis, and tissue patterning.
• Cell Proliferation and Differentiation: FGF-9 stimulates proliferation and differentiation in various cell types, including mesenchymal, nerve, and epithelial cells, which is crucial for in vitro cell culture, organoid formation, and regenerative medicine research.
• Paracrine and Autocrine Signaling: FGF-9 functions both as an autocrine and paracrine factor, influencing local tissue environments and cell-cell communication, especially in prostate and neural tissues.
• Disease Modeling: FGF-9 is implicated in cancer biology, fibrosis, and tissue repair. For example, recombinant FGF-9 can be used to study tumorigenicity, therapy resistance, and cellular migration in hepatocellular carcinoma models.
• Tissue Engineering and Regeneration: Recombinant FGFs, including FGF-9, are used to promote tissue repair and regeneration in animal models, supporting applications in wound healing, bone repair, and organoid development.

Best practices for using recombinant rat FGF-9:

• Employ in cell culture systems to stimulate proliferation, differentiation, or survival of target cell populations.
• Use in organoid or tissue engineering protocols to mimic developmental signaling environments.
• Apply in disease models to investigate FGF-9’s role in pathogenesis or therapeutic response.

In summary, recombinant rat FGF-9 is a versatile tool for research in developmental biology, neurobiology, tissue engineering, and disease modeling, enabling precise control over FGF-9-mediated signaling pathways in vitro and in vivo.

Recombinant Rat FGF-9 can be used as a standard for quantification or calibration in ELISA assays, provided it is compatible with your specific assay system and matches the form of FGF-9 detected by your antibodies.

Key considerations and supporting details:

• Recombinant proteins are commonly used as ELISA standards when they are of high purity and their concentration is accurately known. Many commercial ELISA kits for FGF-9 use recombinant rat FGF-9 as the calibrator or standard to generate the standard curve for quantification.

• Assay compatibility is critical: The recombinant FGF-9 must be recognized by the capture and detection antibodies in your ELISA. Most sandwich ELISAs for rat FGF-9 are designed to detect both natural and recombinant forms, provided the recombinant protein contains the relevant epitopes. However, some kits or antibodies may have reduced affinity for recombinant proteins expressed in certain systems (e.g., E. coli vs. mammalian cells) due to differences in folding or post-translational modifications.

• Standard preparation: The recombinant FGF-9 should be reconstituted and diluted according to best practices, ideally in the same buffer as your samples, to minimize matrix effects. The concentration should be verified, and serial dilutions should be prepared to cover the expected range of your assay.

• Validation: It is recommended to validate the use of your recombinant FGF-9 as a standard by running a standard curve and confirming parallelism with endogenous FGF-9 in your sample matrix. This ensures that the recombinant standard behaves similarly to the native protein in your assay conditions.

• Exceptions: Some ELISA kits specifically advise against using recombinant proteins as standards if their antibodies are optimized only for the native form or if the recombinant protein lacks certain modifications. Always consult your ELISA kit manual or antibody datasheet for such restrictions.

Summary of best practices:

• Confirm that your ELISA antibodies detect recombinant rat FGF-9.
• Prepare the standard curve using accurately quantified recombinant protein.
• Validate parallelism between the standard curve and sample dilution curves.
• Use the same buffer for standards and samples to minimize matrix effects.

If these conditions are met, recombinant rat FGF-9 is suitable as a standard for quantification or calibration in ELISA assays.

Recombinant Rat FGF-9 has been validated for several key applications in published research, primarily in studies involving cell proliferation, differentiation, tissue regeneration, and angiogenesis.

Validated Applications:

• Angiogenesis and Bone Regeneration: Recombinant rat FGF-9 has been shown to enhance angiogenesis, osteogenesis, and bone remodeling in vivo. Application in tibial bone defect models demonstrated increased cell proliferation and differentiation, particularly in osteoblasts, as assessed by immunohistochemistry for markers such as PCNA, RUNX-2, and Osteocalcin.

• Neurobiology and Neural Differentiation: FGF-9 is highly expressed in the brain and functions as a neurotrophic factor. It supports the growth and survival of motor neurons and has been used to study glial activation and neural progenitor cell differentiation.

• Cell Proliferation and Migration Assays: Recombinant FGF-9 has been used in bioassays to stimulate proliferation and migration in various cell types, including hepatic stellate cells and cancer cell lines. It induces ERK and JNK pathway activation, promoting clonogenicity and migration, and has been implicated in therapy resistance in hepatocellular carcinoma models.

• In Vivo Models: FGF-9 has been applied in animal models to study tissue repair, wound healing, and regeneration, including bone and neural tissues. These studies often use whole cells or tissue samples and assess outcomes such as revascularization, cell proliferation, and differentiation.

• Stem Cell and Organotypic Culture: FGF-9 is used to direct differentiation in organoid cultures, particularly for ureteric bud and collecting duct organoids derived from pluripotent stem cells (though this is more commonly reported for human and mouse FGF-9).

Experimental Techniques:

• Bioassays: Used to measure cell proliferation, differentiation, and migration in response to FGF-9 stimulation.
• Immunohistochemistry: For detection of proliferation and differentiation markers in tissue sections.
• In Vivo Regeneration Models: Application in rat models for bone and neural tissue regeneration.
• Cell Culture: Used as a supplement to support growth and survival of specific cell types, including neurons and prostate tissue.

Summary Table:

Key Insights:

• Recombinant rat FGF-9 is most frequently validated for use in bioassays, immunohistochemistry, and in vivo regeneration models.
• Its roles span angiogenesis, osteogenesis, neural survival, and cell proliferation/migration.
• Applications are supported by both in vitro and in vivo studies, with a focus on tissue repair and regeneration.

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

To reconstitute and prepare Recombinant Rat FGF-9 protein for cell culture experiments, follow these best-practice steps:

1. Centrifuge the vial briefly before opening to ensure all lyophilized protein is at the bottom.
2. Reconstitution:
   • Add sterile distilled water or aqueous buffer (such as PBS) to achieve a concentration of at least 100 μg/mL (0.1 mg/mL) or as specified by your protocol.
   • For enhanced stability and to prevent adsorption, it is recommended to include 0.1% carrier protein (such as BSA or HSA) in the buffer.
   • Gently pipette up and down or swirl to dissolve. Do not vortex; allow several minutes for complete dissolution.
3. Aliquot the reconstituted protein to avoid repeated freeze-thaw cycles.
4. Storage:
   • Store aliquots at –20°C to –80°C for long-term storage.
   • For short-term use, store at 2–8°C for up to 1 week.
   • If stored long-term, always include a carrier protein to maintain stability.
5. Further dilution: Before adding to cell culture, dilute the stock solution to the desired working concentration using cell culture medium or appropriate buffer, maintaining the presence of carrier protein if possible.

Summary of key steps:

• Centrifuge vial before opening.
• Reconstitute in sterile water or buffer (≥100 μg/mL), with 0.1% BSA/HSA.
• Gently mix; do not vortex.
• Aliquot and store at –20°C to –80°C.
• Dilute to working concentration in cell culture medium before use.

Additional notes:

• Always consult the specific product datasheet for any unique instructions.
• Avoid repeated freeze-thaw cycles to preserve protein activity.
• For cell culture, ensure all solutions are sterile and endotoxin-free.

These guidelines are consistent with standard protocols for recombinant FGF-9 and other growth factors used in cell culture.