Recombinant Mouse SDF-1α

Recombinant Mouse SDF-1α (also known as CXCL12) is widely used in research because it is a potent chemokine that regulates cell migration, stem cell homing, tissue regeneration, and immune cell trafficking, making it essential for studies in immunology, regenerative medicine, oncology, and developmental biology.

Key scientific applications and rationale include:

• Stem Cell Homing and Migration: SDF-1α is a critical factor for the homing and mobilization of hematopoietic stem cells and other progenitor cells to specific tissues, such as bone marrow and sites of injury. This property is exploited in assays studying stem cell recruitment, tissue repair, and regeneration.

• Tissue Regeneration and Repair: SDF-1α enhances recruitment of progenitor cells to damaged tissues, promoting regeneration in models of cartilage repair, myocardial infarction, and wound healing. For example, exogenous SDF-1α significantly improves cartilage integration and mechanical properties in repair models, and accelerates wound closure and angiogenesis in skin injury models.

• Immunology and Inflammation: SDF-1α recruits activated leukocytes via the CXCR4 receptor, playing a role in immune surveillance, inflammation, and autoimmune disease models. It is used to study leukocyte trafficking, T cell migration, and the formation of pre-metastatic niches in cancer research.

• Developmental Biology: Mouse SDF-1α is essential for normal development of the hematopoietic, cardiovascular, and nervous systems, making it valuable for developmental studies.

• In Vitro and In Vivo Assays: Recombinant SDF-1α is used in migration and homing assays, angiogenesis assays, and bioassays to study cell signaling, proliferation, and chemotaxis.

• Species Specificity: Using recombinant mouse SDF-1α ensures compatibility with mouse models, minimizing cross-species variability and maximizing physiological relevance in murine systems.

In summary, recombinant mouse SDF-1α is a versatile tool for dissecting mechanisms of cell migration, tissue repair, immune response, and development in mouse-based research models, supporting both basic and translational studies.

Yes, recombinant mouse SDF-1α can be used as a standard for quantification or calibration in ELISA assays, provided it is of high purity and its concentration is accurately known. This approach is standard practice in quantitative ELISA protocols for cytokines and chemokines, including SDF-1α/CXCL12.

Key considerations and supporting details:

• Recombinant SDF-1α is commonly used as a standard in commercial mouse SDF-1α ELISA kits. These kits typically use E. coli-expressed recombinant mouse SDF-1α as the reference standard to generate the calibration curve.
• Validation: The ELISA standard curve generated with recombinant SDF-1α is shown to be parallel to curves obtained with natural SDF-1α, indicating that quantification is accurate for both recombinant and natural forms.
• Purity and quantification: Ensure that the recombinant SDF-1α you use is highly purified and that its concentration has been determined accurately (e.g., by absorbance at 280 nm or amino acid analysis). Impurities or inaccurate concentration will compromise assay accuracy.
• Matrix effects: If you are preparing your own standards, dilute the recombinant protein in the same buffer or matrix as your samples (e.g., assay diluent or serum-free media) to minimize matrix effects and ensure comparable binding conditions.
• Bioactivity vs. immunoreactivity: ELISA quantifies immunoreactive SDF-1α, not necessarily bioactive protein. If your application requires functional activity, additional bioassays may be necessary.

Limitations:

• For research use only: Recombinant standards and ELISA kits are generally intended for research, not diagnostic, purposes.
• Isoform specificity: Confirm that the recombinant protein matches the isoform detected by your ELISA antibodies (e.g., SDF-1α vs. SDF-1β), as cross-reactivity may vary.

Protocol tip:
Prepare a serial dilution of the recombinant SDF-1α in the recommended assay diluent to generate a standard curve covering the expected concentration range in your samples.

In summary, using recombinant mouse SDF-1α as a standard is scientifically valid and widely practiced for ELISA quantification, provided you follow best practices for protein standards and assay setup.

Recombinant Mouse SDF-1α (CXCL12) has been validated in published research for a range of applications, primarily related to its role as a chemokine in cell migration, immune modulation, and tissue regeneration.

Key validated applications include:

• Cell migration and chemotaxis assays: SDF-1α is widely used to induce and study the migration of various cell types, including T lymphocytes, monocytes, hematopoietic stem cells, endothelial progenitor cells, and mesenchymal stem cells, often via the CXCR4 receptor.
• Wound healing and tissue regeneration models: Recombinant SDF-1α has been applied in murine models to enhance wound healing, particularly by promoting angiogenesis and recruiting progenitor cells to sites of injury.
• Stem cell homing and engraftment: It is used to study and enhance the homing of hematopoietic and mesenchymal stem cells to bone marrow and injured tissues, both in vitro and in vivo.
• Bioassays for functional activity: SDF-1α is validated in bioassays measuring chemotactic responses, receptor activation, and downstream signaling events in various cell types.
• Immunological studies: It is used to investigate immune cell trafficking, inflammation, and autoimmune disease mechanisms, including models of lupus and experimental autoimmune encephalomyelitis.
• Cancer research: SDF-1α is employed in studies of tumor microenvironment, metastasis, and the recruitment of tumor-associated cells, such as macrophages and endothelial progenitors.
• Cell culture supplementation: It is used to support cell survival, proliferation, and differentiation in vitro, particularly for hematopoietic and neural cells.

Additional validated uses include:

• Angiogenesis assays (e.g., tube formation, endothelial cell outgrowth).
• Adhesion assays to study cell–matrix and cell–cell interactions.
• Neuronal development studies and organogenesis models.

These applications are supported by numerous peer-reviewed publications and product validation data, with SDF-1α being a standard reagent for chemotaxis, stem cell biology, regenerative medicine, and immunology research.

To reconstitute and prepare Recombinant Mouse SDF-1α (CXCL12) protein for cell culture experiments, follow these steps for optimal solubility, stability, and biological activity:

1. Centrifuge the vial briefly before opening to ensure all lyophilized protein is at the bottom.

2. Reconstitution:

   • Add sterile distilled water or sterile PBS to the vial to achieve a final concentration between 0.1–1.0 mg/mL.
   • If recommended by the datasheet, include a carrier protein such as 0.1–1% BSA (bovine serum albumin) or heat-inactivated fetal calf serum to stabilize the protein and minimize adsorption to tube walls.
   • Gently pipette the solution down the sides of the vial to dissolve the protein. Do not vortex to avoid denaturation.

3. Incubation:

   • Allow the protein to dissolve at room temperature for 15–30 minutes with gentle agitation.
   • Ensure complete dissolution before proceeding.

4. Aliquoting and Storage:

   • Prepare working aliquots to avoid repeated freeze-thaw cycles.
   • Store aliquots at 2–8°C for up to one week or at –20°C to –80°C for long-term storage.
   • For extended storage, dilute in buffer containing 0.1% BSA.

5. Preparation for Cell Culture:

   • Make further dilutions in tissue culture-grade buffered solutions (e.g., PBS or cell culture medium) containing low endotoxin BSA or heat-inactivated serum.
   • Typical working concentrations for cell assays range from 10–100 ng/mL, but optimal concentrations should be determined empirically for your specific application.

Best Practices:

• Always consult the specific product datasheet for recommended reconstitution buffer and concentration, as formulations may vary.
• Avoid vigorous mixing or vortexing to prevent protein denaturation.
• Use low endotoxin reagents and maintain sterile technique throughout preparation to prevent contamination and ensure cell viability.

Summary Table: Recombinant Mouse SDF-1α Reconstitution

This protocol ensures maximum solubility, stability, and biological activity of recombinant SDF-1α for cell culture applications.