Recombinant Rat Jagged-1

Recombinant Rat Jagged-1 is used in research applications to study and manipulate Notch signaling, a pathway critical for cell fate decisions, differentiation, proliferation, and tissue regeneration. Its recombinant form allows precise, reproducible activation or inhibition of Notch pathways in controlled experimental settings.

Key scientific applications and rationale include:

• Stem Cell Biology: Jagged-1 enhances the survival and expansion of stem cells, particularly hematopoietic and mesenchymal stem cells, and can augment their functional capacity for tissue regeneration and reconstitution. This is valuable for studies on stem cell maintenance, differentiation, and transplantation.

• Developmental Biology: Jagged-1 is essential for processes such as hematopoiesis, myogenesis, neurogenesis, and vascular development. Recombinant protein enables investigation of these pathways in vitro and in vivo, including lineage specification and organogenesis.

• Tissue Engineering and Regeneration: Recombinant Jagged-1 has been shown to promote healing and regeneration of bone and other tissues, making it useful for regenerative medicine models and biomaterial engineering. For example, intraoperative delivery of Jagged-1 accelerates bone defect repair in animal models.

• Cancer and Disease Models: Jagged-1 is implicated in tumor progression, immune evasion, and disease states such as osteoarthritis and diabetic nephropathy. Recombinant protein allows mechanistic studies of these processes and testing of therapeutic interventions.

• Cell Differentiation and Fate Decisions: By binding to Notch receptors, recombinant Jagged-1 can direct cell fate, such as promoting endothelial or neural differentiation, or inhibiting epithelial-mesenchymal transition (EMT).

• Bioassays and ELISA Standards: Recombinant Jagged-1 is used as a standard or ligand in bioassays, ELISA, and cell culture experiments to quantify or modulate Notch pathway activity.

Best practices:

• Use carrier-free formulations for bioassays and mechanistic studies to avoid interference from stabilizing agents.
• Select appropriate concentrations and controls to ensure specificity of Notch pathway activation.

Summary:
Recombinant Rat Jagged-1 is a versatile tool for dissecting Notch signaling in diverse biological contexts, enabling controlled studies of cell fate, tissue regeneration, disease mechanisms, and therapeutic development.

You can use recombinant rat Jagged-1 as a standard for quantification or calibration in your ELISA assays, provided it is properly validated and matched to your assay system. This is a common and accepted practice in ELISA development and quantification, as long as the recombinant protein is of high purity and its concentration is accurately known.

Key considerations and supporting details:

• Recombinant protein as standard: ELISA kits and protocols routinely use purified recombinant proteins as standards to generate calibration curves for quantification. This is specifically mentioned in technical guidelines for ELISA standard preparation.
• Assay compatibility: Many commercial rat Jagged-1 ELISA kits are designed to detect both natural and recombinant rat Jagged-1, and their standard curves are often generated using recombinant protein. This indicates that recombinant rat Jagged-1 is suitable for use as a standard, assuming your assay antibodies recognize the recombinant form equivalently to the native protein.
• Validation: It is essential to validate that your recombinant Jagged-1 standard behaves similarly to the endogenous protein in your assay matrix (e.g., serum, plasma, cell culture supernatant). This includes confirming parallelism between the standard curve and sample dilution curves, and ensuring there is no matrix interference.
• Concentration accuracy: The recombinant protein must be accurately quantified (e.g., by absorbance at 280 nm, BCA assay, or amino acid analysis) to ensure the standard curve reflects true concentrations.
• Standard curve range: Ensure the concentration range of your recombinant standard matches the expected range of Jagged-1 in your samples and the dynamic range of your ELISA (e.g., 73–18,000 pg/mL in some kits).

Best practices:

• Prepare serial dilutions of the recombinant Jagged-1 in the same buffer or matrix as your samples to minimize matrix effects.
• Validate the standard curve for linearity, precision, and accuracy within your assay system.
• If using a kit, follow the manufacturer’s recommendations regarding standard preparation and compatibility with recombinant proteins.

In summary: Recombinant rat Jagged-1 is appropriate for use as a standard in ELISA quantification, provided you validate its performance in your specific assay context and ensure accurate concentration determination.

Recombinant Rat Jagged-1 has been validated in published research primarily for bioassays involving Notch signaling modulation, cell differentiation, and tissue regeneration, with applications in both in vitro and in vivo models.

Key validated applications include:

• Bioassays: Used to activate or inhibit Notch signaling in cultured cells, including studies on stem cell fate, endothelial differentiation, and immune cell function.
• Cell differentiation studies: Demonstrated to induce differentiation of mesenchymal stem cells into endothelial cells, and to influence fate decisions in cartilage progenitor/stem cells.
• Cancer research: Employed in rat xenograft models to study tumor growth, immune evasion, and the effects of JAG1 blockade on tumor progression and the tumor microenvironment.
• Tissue regeneration and repair: Used in models of bone regeneration and healing, including delivery via hydrogels to stimulate bone repair in vivo.
• In vivo functional studies: Applied in rat models to investigate roles in intestinal adaptation after small bowel resection and in muscle regeneration.
• ELISA and Western blot: Validated as a standard or control protein for immunoassays detecting Jagged-1 or related pathway components.

Sample types validated include:

• Whole cells (primary cells, cell lines)
• Organoids
• Tissue explants
• In vivo rat models (e.g., xenografts, tissue injury models)

Summary of published research applications:

These applications demonstrate that recombinant rat Jagged-1 is a versatile tool for dissecting Notch pathway biology, cell fate decisions, and tissue regeneration in both basic and translational research contexts.

To reconstitute and prepare Recombinant Rat Jagged-1 protein for cell culture experiments, follow these general guidelines based on manufacturer protocols and best practices:

1. Reconstitution

• Lyophilized Protein: Most recombinant Rat Jagged-1 proteins (e.g., Fc Chimera or His-tagged) are supplied lyophilized.
• Reconstitution Buffer: Reconstitute the protein in sterile PBS (phosphate-buffered saline) or as specified in the product datasheet. For example:
  • Recombinant Rat Jagged-1 Fc Chimera: Reconstitute at 200 μg/mL in sterile PBS.
  • Recombinant Rat Jagged-1 His-tagged: Reconstitute in sterile PBS or as directed.
• Procedure:
  • Centrifuge the vial briefly before opening to ensure all powder is at the bottom.
  • Add the appropriate volume of sterile PBS to achieve the desired concentration.
  • Gently swirl or pipette up and down to dissolve the protein. Avoid vigorous mixing or vortexing to prevent denaturation.
  • Allow the solution to sit for 15–30 minutes at room temperature to ensure complete dissolution.

2. Storage After Reconstitution

• Short-term: Store reconstituted protein at 2–8°C for up to 1 month under sterile conditions.
• Long-term: Aliquot and store at –20°C to –70°C for up to 3 months. Avoid repeated freeze-thaw cycles.

3. Preparation for Cell Culture

• Dilution: Dilute the reconstituted protein in cell culture medium or PBS to the desired working concentration. Optimal concentrations may vary depending on the experiment (e.g., 1–10 μg/mL for coating or soluble ligand applications).
• Sterility: Ensure all solutions and containers are sterile to prevent contamination.
• Application:
  • For coating surfaces: Dilute the protein in PBS and coat plates (e.g., 5–10 μg/mL in 100 μL per well for a 96-well plate). Incubate at 4°C overnight or at 37°C for 2 hours. Wash plates before adding cells.
  • For soluble ligand: Add the diluted protein directly to the culture medium.

4. Best Practices

• Avoid Repeated Freeze-Thaw Cycles: Aliquot the reconstituted protein to minimize degradation.
• Mix Gently: Always mix gently to avoid foaming or denaturation.
• Check Product Datasheet: Always refer to the specific product datasheet for any unique instructions or buffer requirements.

Example Protocol (Fc Chimera Protein):

1. Reconstitute 10 μg of lyophilized protein in 50 μL sterile PBS (200 μg/mL).
2. Mix gently and let sit for 15–30 minutes at room temperature.
3. Aliquot and store at –20°C for long-term use.
4. For cell culture, dilute to working concentration (e.g., 5 μg/mL) in culture medium or PBS.

By following these steps, you can ensure proper reconstitution and preparation of Recombinant Rat Jagged-1 protein for reliable cell culture experiments.