Recombinant Human BMP-13

Recombinant Human BMP-13 (also known as GDF6) is primarily used in research for its unique roles in cartilage and tendon formation, tissue repair, and the regulation of mesenchymal stem cell differentiation, making it valuable for studies in regenerative medicine and tissue engineering.

Key scientific reasons to use recombinant human BMP-13 in research applications:

• Cartilage and Tendon Regeneration: BMP-13 is a potent inducer of chondrogenic (cartilage-forming) and tenogenic (tendon-forming) differentiation. It promotes the formation of cartilage and tendon-like tissues, which is critical for developing therapies for joint, tendon, and ligament injuries.

• Tissue Engineering and Repair: BMP-13 is widely studied for its ability to enhance tissue repair, especially in musculoskeletal tissues. It has been shown to upregulate extracellular matrix proteins such as type III collagen and fibronectin, improving the biomechanical properties of repaired tissues.

• Regulation of Mesenchymal Stem Cell Fate: Unlike other BMPs (such as BMP-2 and BMP-7), BMP-13 inhibits osteogenic (bone-forming) differentiation of bone marrow-derived mesenchymal stem cells (BM MSCs) while promoting chondrogenic differentiation. This makes it particularly useful when the goal is to prevent unwanted bone formation and instead encourage cartilage or tendon regeneration.

• Maintenance of Articular Cartilage: BMP-13 is expressed in articular cartilage postnatally and is thought to play a regulatory role in the growth and maintenance of cartilage, making it relevant for studies on osteoarthritis and cartilage repair.

• Modeling Disease and Development: BMP-13 is implicated in skeletal development and certain congenital disorders (e.g., Klippel-Feil syndrome), so it is useful for modeling these conditions in vitro and in vivo.

Summary of applications:

• Promoting cartilage and tendon repair in tissue engineering models.
• Studying the molecular mechanisms of chondrogenesis and tenogenesis.
• Investigating the regulation of stem cell differentiation away from osteogenesis and toward soft tissue lineages.
• Exploring therapeutic strategies for degenerative joint diseases and soft tissue injuries.

Distinct from other BMPs:
BMP-13 is less osteo-inductive than BMP-2 or BMP-7, so it is preferred when bone formation is not desired or may be detrimental to the research goal.

In summary, recombinant human BMP-13 is a specialized tool for research focused on cartilage, tendon, and ligament regeneration, as well as for dissecting the pathways that control mesenchymal stem cell fate in musculoskeletal tissues.

Yes, recombinant human BMP-13 can be used as a standard for quantification and calibration in ELISA assays, though there are important considerations regarding formulation and intended use.

Suitability as an ELISA Standard

Recombinant BMP-13 is appropriate for ELISA standardization when properly prepared. For standard curve development, a purified protein should ideally be used, or alternatively, a recombinant protein that can be semi-purified in the laboratory and measured with high-performance liquid chromatography (HPLC). Recombinant BMP-13 meets these criteria as a suitable calibration material.

Protein Characteristics

Recombinant human BMP-13 exists as a 27.0 kDa homodimeric disulfide-linked protein consisting of two 120 amino acid polypeptide chains. This structural information is important for understanding its behavior in immunoassays and ensuring proper reconstitution and handling.

Standard Curve Parameters

When using recombinant BMP-13 as an ELISA standard, typical standard curve ranges extend from 0.16 to 10 ng/mL, with sensitivity around 0.09 ng/mL. More generally, standard curves typically range from 0 to 1000 pg/mL, though this can extend to 3000 pg/mL if target protein concentrations are predicted to be extremely high.

Important Formulation Considerations

A critical distinction exists between ELISA standard formulations and bioassay-grade recombinant proteins. ELISA standard recombinant proteins are specifically formulated and validated for quantification purposes but are not recommended for bioassay applications, as they have not been tested for biological activity. Conversely, if you require functional assessment of BMP-13 activity, you would need bioassay-grade material rather than ELISA standard formulations.

For cell or tissue culture applications combined with ELISA standardization, recombinant BMP-13 formulated with bovine serum albumin (BSA) as a carrier protein is typically recommended.

Reconstitution and Storage

Most recombinant protein standards are provided in lyophilized form and require reconstitution according to manufacturer-specific instructions, as protocols may be lot-specific. Proper storage conditions must be maintained to preserve protein integrity and ensure reliable quantification results.

Recombinant Human BMP-13 (also known as GDF6 or CDMP-2) has been validated in published research for several key applications, primarily in the fields of connective tissue engineering, stem cell differentiation, and inhibition of osteogenesis.

Validated Applications in Published Research:

• Induction of Tendon and Ligament Differentiation:
  Recombinant human BMP-13 has been shown to induce differentiation of mesenchymal stem cells (MSCs) and anterior cruciate ligament (ACL) fibroblasts into ligament-like cells when cultured in a type I collagen hydrogel. It also upregulates type III collagen and fibronectin expression and improves biomechanical properties in tendon repair models.

• Chondrogenic Differentiation:
  BMP-13 can induce chondrogenic phenotypes, characterized by increased proteoglycan production in human fetal chondrocytes and mouse chondrocytic cell lines. This supports its use in cartilage tissue engineering and repair.

• Inhibition of Osteogenic Differentiation:
  BMP-13 has been validated as an inhibitor of osteogenic differentiation in human bone marrow-derived mesenchymal stromal cells (BM MSCs) in vitro. It dose-dependently reduces alkaline phosphatase (ALP) activity and mineralization, distinguishing it from other BMPs like BMP-2 and BMP-7, which promote bone formation.

• Augmentation of Osteochondral Repair:
  Adenovirus-mediated BMP-13 gene transfer to rabbit bone marrow stem cells has been reported to enhance periosteal repair of osteochondral defects, indicating its application in joint and cartilage repair models.

• Functional and Blocking Assays:
  Recombinant BMP-13 (GDF6) has been validated for use in functional assays (e.g., induction of ALP in chondrogenic cell lines) and blocking assays, supporting its utility in mechanistic studies of signaling pathways.

Summary Table of Validated Applications

Additional Notes:

• BMP-13 is not strongly osteo-inductive compared to BMP-2 or BMP-7, making it more suitable for applications where inhibition of bone formation or promotion of soft tissue (tendon/ligament/cartilage) regeneration is desired.
• Most published research focuses on in vitro and preclinical in vivo models; clinical validation is limited.

These applications are supported by peer-reviewed studies and reviews, reflecting the current state of research on recombinant human BMP-13.

To reconstitute and prepare Recombinant Human BMP-13 for cell culture experiments, dissolve the lyophilized protein in sterile water or buffer to a concentration of at least 100 µg/mL, then dilute as needed for your assay using a carrier protein-containing solution to prevent loss of activity and adsorption to surfaces.

Step-by-step protocol:

1. Centrifuge the vial: Before opening, briefly centrifuge the vial to collect all lyophilized powder at the bottom.

2. Reconstitution:

   • Add sterile, ultrapure water (18 MΩ·cm) or the recommended buffer directly to the vial to achieve a concentration of ≥100 µg/mL.
   • Gently mix by swirling or inverting; do not vortex or shake vigorously to avoid protein denaturation.
   • Allow the protein to fully dissolve (typically a few minutes at room temperature).

3. Dilution for cell culture:

   • For working concentrations, dilute the reconstituted BMP-13 in cell culture medium or a buffer containing a carrier protein (e.g., 0.1% BSA, 10% FBS, or 5% HSA) to minimize adsorption and maintain activity, especially at low concentrations.
   • If using serum-free conditions, use a non-animal stabilizer such as trehalose.

4. Storage:

   • Short-term (2–7 days): Store reconstituted BMP-13 at 2–8 °C.
   • Long-term: For storage beyond one week, aliquot and freeze at ≤–20 °C (preferably –80 °C) with a carrier protein or 5–50% glycerol to prevent freeze-thaw damage.
   • Avoid repeated freeze-thaw cycles by aliquoting.

5. Handling tips:

   • Do not exceed 1 mg/mL during reconstitution to avoid solubility issues.
   • Confirm protein integrity by SDS-PAGE if needed.
   • Always consult the product-specific datasheet or Certificate of Analysis for any unique requirements.

Summary Table: BMP-13 Reconstitution and Storage

Note: Always verify the specific requirements for your BMP-13 preparation, as formulation and recommended buffers may vary between products. If using for in vivo or serum-free applications, avoid animal-derived carriers and use alternatives like trehalose.