Recombinant Human FGF-19

Recombinant Human FGF-19 is used in research applications because it is a potent regulator of metabolism, muscle mass, and liver function, with demonstrated benefits in models of metabolic disease, muscle wasting, and tissue regeneration.

FGF-19 is an endocrine fibroblast growth factor that acts primarily through the FGFR4/β-Klotho receptor complex, influencing multiple physiological processes:

• Metabolic Regulation: FGF-19 lowers blood glucose, reduces hepatic fat, and decreases plasma bile acid and cholesterol levels, making it valuable for studying diabetes, non-alcoholic fatty liver disease (NAFLD), and related metabolic disorders.
• Muscle Biology: Recombinant FGF-19 increases skeletal muscle fiber size, improves muscle strength, and counteracts muscle wasting (sarcopenia) in preclinical models, including chronic kidney disease (CKD) and aging. These effects are mediated via the ERK/mTOR signaling pathway and are independent of food intake or weight gain.
• Liver and Bile Acid Homeostasis: FGF-19 regulates hepatic bile acid synthesis and supports liver regeneration, making it useful for research on cholestasis, liver injury, and hepatic proliferation.
• Anti-inflammatory and Anti-fibrotic Effects: FGF-19 treatment reduces hepatic expression of inflammatory markers and genes involved in gluconeogenesis, supporting its use in studies of liver inflammation and metabolic stress.
• Cancer and Cell Proliferation: FGF-19 is implicated in cancer biology, particularly in promoting proliferation in certain tumor types, so it is used to study oncogenic signaling pathways and potential therapeutic interventions.

Common research applications include:

• In vitro bioassays to study receptor signaling and downstream effects.
• Organoid and cell culture models for liver, muscle, and metabolic disease research.
• In vivo studies to evaluate therapeutic potential in metabolic, muscular, and hepatic disorders.
• Investigating tissue regeneration and repair, given FGF-19’s role in cell proliferation and differentiation.

In summary, recombinant human FGF-19 is a versatile tool for dissecting metabolic pathways, muscle and liver physiology, and disease mechanisms, as well as for preclinical evaluation of novel therapeutic strategies targeting metabolic and degenerative diseases.

Recombinant human FGF-19 can be used as a standard for quantification or calibration in ELISA assays, provided it is validated for this purpose and matches the assay’s requirements.

Key considerations and supporting details:

• ELISA Standards: Commercial FGF-19 ELISA kits typically use recombinant human FGF-19 as the standard for generating calibration curves. These standards are usually E. coli-expressed or mammalian cell-expressed recombinant proteins, and their concentrations are carefully determined and validated for use in the specific ELISA format.

• Validation and Parallelism: For accurate quantification, the recombinant FGF-19 standard must demonstrate parallelism with endogenous (natural) FGF-19 in your sample matrix. This means the standard curve generated with recombinant FGF-19 should be parallel to the dilution curves of your biological samples, indicating similar antibody recognition and assay response. Some kits specifically state that their recombinant standard and natural FGF-19 yield parallel curves, supporting their use for quantification.

• Source and Purity: The recombinant FGF-19 used as a standard should be of high purity, with a well-characterized concentration, and free from contaminants such as endotoxins or carrier proteins that could interfere with the assay. The protein should be stored and handled according to best practices to maintain stability and activity.

• Assay Compatibility: Not all recombinant FGF-19 preparations are suitable for use as ELISA standards. Some recombinant proteins are intended for bioassays or functional studies and may not be validated for use as ELISA standards. Always confirm that the recombinant FGF-19 you intend to use is suitable for ELISA calibration, ideally by referencing the manufacturer’s documentation or by performing a validation experiment.

• Best Practices:

  • Prepare a standard curve using serial dilutions of the recombinant FGF-19 in the same buffer or matrix as your samples.
  • Confirm that the standard curve covers the expected concentration range of FGF-19 in your samples.
  • Validate recovery and linearity by spiking known amounts of recombinant FGF-19 into your sample matrix and assessing recovery rates.

Summary Table: Recombinant FGF-19 as ELISA Standard

In conclusion:
You can use recombinant human FGF-19 as a standard for ELISA quantification if it is validated for this purpose, matches the assay’s requirements, and demonstrates parallelism with endogenous FGF-19 in your sample matrix. Always verify compatibility and perform appropriate controls to ensure accurate quantification.

Recombinant Human FGF-19 has been validated for a wide range of applications in published research, including:

• Bioassay: Used to study proliferation, migration, and invasion in various cell types, including cancer cells, organoids, and primary hepatocytes.
• Organoid Culture: Employed in the expansion and maintenance of human and porcine liver organoids.
• Functional Assay: Applied in studies assessing the effects of FGF-19 on muscle fiber size, muscle strength, and metabolic parameters in animal models.
• ELISA: Used as a standard or for detection in immunoassays.
• Immunohistochemistry (IHC): Validated for detecting FGF-19 and its effects in tissue sections.
• Western Blot (WB): Used to analyze protein expression and signaling pathways.
• Surface Plasmon Resonance (SPR): Applied in studies investigating protein-protein interactions.
• Blocking Assay: Used to study the inhibition of FGF-19 signaling.
• Neuroprotective Studies: Investigated for its effects on adult mammalian neurons.
• Metabolic Studies: Used to assess effects on glucose homeostasis, insulin sensitivity, and hepatic metabolism.

These applications highlight the versatility of Recombinant Human FGF-19 in both basic research and preclinical studies.

To reconstitute and prepare Recombinant Human FGF-19 protein for cell culture experiments, follow these best-practice steps:

1. Centrifuge the vial briefly (e.g., 30 seconds at 3,000 rpm) before opening to ensure all lyophilized protein is at the bottom.

2. Reconstitution buffer:

   • Use sterile 1× PBS (pH 7.2–7.4) as the primary buffer.
   • For enhanced stability and to prevent adsorption, add a carrier protein such as 0.1%–1% endotoxin-free human or bovine serum albumin (HSA/BSA).
   • Alternatively, some protocols allow reconstitution in sterile distilled water, but PBS with carrier protein is preferred for cell culture applications.

3. Protein concentration:

   • Common stock concentrations are 100 μg/mL or 0.2 mg/mL.
   • Adjust the volume of buffer accordingly to achieve the desired concentration.

4. Mixing:

   • Gently swirl or tap the vial to dissolve the protein. Avoid vigorous vortexing, which can denature the protein.

5. Aliquoting and storage:

   • After reconstitution, aliquot the solution to avoid repeated freeze-thaw cycles.
   • Store aliquots at –20°C to –80°C for long-term storage (up to 6 months), or at 4°C for short-term use (up to 1 week).
   • Avoid frost-free freezers and repeated freeze-thaw cycles, as these can degrade the protein.

6. Working solution:

   • For cell culture, dilute the stock solution to the desired working concentration using cell culture medium or PBS with carrier protein.
   • Prepare fresh working solutions for each experiment if possible.

Summary Table: FGF-19 Reconstitution and Handling

Additional notes:

• If the protein does not fully dissolve, allow it to sit at room temperature for a few minutes and gently mix again.
• If a precipitate forms, clarify by brief centrifugation before use.
• Always consult the product-specific Certificate of Analysis (CoA) or datasheet for any unique instructions.

These steps will help ensure maximum stability and bioactivity of recombinant FGF-19 for your cell culture experiments.