Recombinant Human SCF

Recombinant human stem cell factor (SCF) is a versatile cytokine with multiple compelling applications across hematopoietic research, cell differentiation studies, and regenerative medicine. Here are the key reasons to incorporate it into your research:

Hematopoietic Applications

SCF is a primary growth and activation factor for hematopoietic stem cells and multiple progenitor populations. It stimulates proliferation and expansion of myeloid, erythroid, lymphoid, and megakaryocytic progenitors in bone marrow cultures. The protein acts through the c-Kit/CD117 receptor, which transmits SCF signals at different levels of both embryonic and adult hematopoiesis. This makes it essential for in vitro expansion of CD34+ hematopoietic progenitor cells and differentiation of embryonic stem cell-derived cells toward the hematopoietic lineage.

Mast Cell and Immune Cell Development

SCF plays a critical role in mast cell differentiation, maintenance, and chemotaxis assays. Beyond mast cells, recombinant SCF is used to generate T cells for cell-based therapies, drug screening, and disease modeling. Additionally, it can be employed in combination with other cytokines to generate myeloid-derived suppressor cells from human umbilical cord blood for treating graft versus host disease.

Stem Cell Maintenance and Differentiation

SCF maintains stemness in various cell populations, including human bone marrow mesenchymal cells and hematopoietic stem cells within their niche environment. The protein is also versatile in differentiating specific cell types such as spermatogonial stem cells and megakaryocyte progenitors. It has improved protocols for continuous generation of cells in culture systems, including granulocytes and macrophages.

Regenerative Medicine and Tissue Engineering

In regenerative applications, SCF is incorporated into wound healing hydrogels to increase adhesion strength and promote tissue regeneration. Preclinical studies have demonstrated that recombinant SCF can protect against lethal irradiation and elicit multilineage hematopoietic responses with increases in bone marrow recovery.

Practical Research Advantages

Recombinant SCF offers high lot-to-lot consistency and enables seamless transition from basic research to clinical applications. The protein is available in multiple formulations—including carrier-free versions for applications where bovine serum albumin would interfere, and animal-free grades for clinical-grade work. Bioactivity is typically measured at ED₅₀ values of 1.00-8.00 ng/mL in cell proliferation assays, providing reliable dose-response relationships for experimental design.

Human-cell-expressed recombinant SCF variants offer additional advantages including native glycosylation, superior stability in culture, and high biological activity compared to prokaryotic expression systems, making them particularly suitable for demanding clinical and therapeutic applications.

You can use recombinant human SCF as a standard for quantification or calibration in your ELISA assays, provided it is of high purity and its concentration is accurately determined. This is a common and accepted practice in ELISA development and quantification protocols.

Key considerations and supporting details:

• Recombinant SCF as Standard: Recombinant human SCF, especially when expressed in E. coli and purified, is routinely used as a standard in commercial SCF ELISA kits. These kits calibrate their standard curves using recombinant SCF, and results for natural samples are reported to be parallel to those obtained with the recombinant standard, indicating suitability for quantification.

• Purity and Quantification: For accurate calibration, the recombinant SCF should be highly purified, and its concentration should be determined using a reliable method (e.g., HPLC, absorbance at 280 nm with a known extinction coefficient). Carrier proteins like BSA are sometimes added for stability, but carrier-free forms are also available and may be preferable if BSA could interfere with your assay.

• Parallelism and Recovery: Commercial kits report that standard curves generated with recombinant SCF are parallel to those obtained with endogenous SCF in biological samples, supporting its use for quantification. Recovery experiments in various matrices (serum, plasma, cell culture media) show high accuracy, further validating this approach.

• Best Practices:

  • Reconstitute the recombinant SCF according to the manufacturer’s instructions, as lot-specific variations can affect concentration.
  • Prepare a fresh standard curve for each assay to ensure quantitative accuracy.
  • If using a recombinant SCF not specifically validated for ELISA, confirm that it is recognized by the antibodies in your assay and that it produces a standard curve with appropriate linearity and parallelism to native SCF.

• Limitations: If your recombinant SCF differs significantly in structure (e.g., glycosylation, isoform) from the endogenous protein in your samples, this could affect antibody recognition and quantification accuracy. Most commercial recombinant SCF standards are the soluble form (e.g., KL-1 variant, 165 aa).

Summary Table: Recombinant SCF as ELISA Standard

In summary, recombinant human SCF is suitable as a standard for ELISA quantification, provided it is well-characterized and compatible with your assay system.

Recombinant Human SCF (Stem Cell Factor) has been validated for a broad range of applications in published research, primarily in cell biology, hematopoiesis, stem cell research, immunology, and regenerative medicine.

Key validated applications include:

• Cell Proliferation and Survival Assays: SCF stimulates proliferation of hematopoietic progenitor cells (e.g., TF-1 erythroleukemic cells), mast cells, and other lineages in bioassays.
• Stem Cell Expansion and Differentiation: Widely used for in vitro expansion of CD34+ hematopoietic stem and progenitor cells, differentiation of embryonic stem (ES) cells toward hematopoietic lineages, and maintenance of pluripotent stem cells.
• Mast Cell Biology: Promotes mast cell differentiation, survival, activation, and chemotaxis; used in mast cell functional assays.
• Regenerative Medicine: Incorporated into wound healing hydrogels to enhance tissue regeneration and adhesion strength.
• Immunology and Cell Therapy: Used to generate T cells for cell-based therapies, drug screening, and disease modeling; supports generation of myeloid-derived suppressor cells for graft-versus-host disease protocols.
• Functional Assays: Validated for use in ELISA, Western blot, immunohistochemistry, blocking assays, and SDS-PAGE for protein characterization and quantification.
• Disease Modeling: Applied in protocols for modeling hematopoietic and immune diseases, including studies on STAT3 signaling in cancer and airway inflammation.
• Cell Culture Supplementation: Used to maintain stemness and support continuous generation of granulocytes, macrophages, and other cell types in culture systems.

Additional validated uses:

• Cardiac Regeneration: Shown to assist in recovery of cardiac function post-myocardial infarction by increasing cardiomyocyte and vascular channel numbers.
• Spermatogonial Stem Cell and Megakaryocyte Progenitor Differentiation: Facilitates differentiation of specific cell types in developmental biology studies.
• Pluripotent Stem Cell Research: Supports reprogramming, maintenance, and differentiation workflows for human pluripotent stem cells.

Summary Table of Validated Applications

These applications are supported by published research and product validation data, demonstrating the versatility of recombinant human SCF in both basic and translational biomedical studies.

To reconstitute and prepare Recombinant Human SCF (Stem Cell Factor) protein for cell culture experiments, follow these best-practice steps:

1. Centrifuge the vial before opening to ensure all lyophilized powder is at the bottom and not lost when opening.

2. Reconstitution buffer:

   • For most applications, use sterile water or sterile PBS (phosphate-buffered saline).
   • If the product contains no carrier protein, consider adding 0.1% BSA (bovine serum albumin) or serum albumin to the buffer to stabilize the protein, especially for long-term storage or when working at low concentrations.

3. Concentration:

   • Common reconstitution concentrations are 0.1 mg/mL (100 μg/mL) or up to 1.0 mg/mL depending on your experimental needs and the manufacturer's recommendation.
   • For example, reconstitute at 100 μg/mL in PBS for a 10 μg vial, or at 500 μg/mL for larger vials.

4. Dissolving the protein:

   • Add the calculated volume of buffer directly to the vial.
   • Gently pipette up and down and wash the sides of the vial to ensure complete dissolution.
   • Do not vortex as this can denature the protein.
   • Allow the solution to sit at room temperature for 15–30 minutes with gentle agitation to ensure full reconstitution.

5. Aliquoting and storage:

   • Once fully dissolved, aliquot the solution to avoid repeated freeze-thaw cycles.
   • Store aliquots at –20°C to –80°C for long-term storage, or at 2–8°C for short-term use (up to 1 week).
   • If using carrier-free protein, always add a carrier protein before freezing to prevent loss due to adsorption or degradation.

6. Working solution:

   • Dilute the stock solution to the desired working concentration using cell culture medium or buffer, ideally supplemented with a carrier protein if not already present.
   • For serum-free cultures, avoid animal-derived carrier proteins and consider using trehalose or other non-animal stabilizers.

Summary Table: Key Steps for SCF Reconstitution

Additional notes:

• Always consult the specific product datasheet or Certificate of Analysis for any unique requirements.
• For cell culture, ensure all solutions are sterile and endotoxin-free.
• If using for in vivo or serum-free applications, avoid animal-derived stabilizers and use alternatives like trehalose.

These steps will help ensure optimal activity and stability of recombinant human SCF for your cell culture experiments.