In Vitro Osteoclast 10-Day Rapid Protocol: M-CSF 30 ng/mL + RANKL 50 ng/mL, TRAP⁺ Multinucleated ≥90%, Bone Resorption Pit Area >70%

One-Sentence Conclusion

Continuous induction of CD14⁺ monocytes with UA-M-CSF (30 ng/mL) and UA-RANKL (50 ng/mL) for 10 days yields ≥90% TRAP⁺ multinucleated osteoclasts and >70% bone resorption pit area, meeting the requirements for osteoporosis drug efficacy evaluation and gene function studies.

The "Dual-Signal" Model for Osteoclast Differentiation

• M-CSF: Survival + Proliferation + Upregulation of RANK receptor (UA040016).
  
  

• RANKL: Fusion + Bone resorption function switch (UA040336).
  
  

• Differentiation fails if either is absent; multinucleation rate <30% if concentrations are insufficient.
  

4-Step Standard Protocol

Gold Standards for Quality Control

• TRAP Staining: Positive rate ≥90% (purple-red cytoplasm + multinucleated).
  
  

• Bone Resorption Assay (e.g., on Osteo Assay Surface): Pit area ≥70%, pit depth ≥3 μm (confirmed by SEM).
  
  

• Genetic Markers: >20-fold upregulation of CTSK, NFATc1, DC-STAMP expression (qPCR).
  

3 Major Advantages of UA Protein Cytokines

• High Activity: RANKL EC₅₀ ≤2 ng/mL, 1.5x higher than leading international brands.
  
  

• Low Endotoxin: <0.05 EU/μg, suitable for direct use in high-throughput drug screening.
  
  

• Batch Consistency: CV <5% across 20 consecutive batches, ensuring long-term experimental reproducibility.
  

5 Major Application Scenarios

• IC₅₀ determination of bone resorption inhibition rate for novel osteoporosis drugs.
  
  

• Functional validation of gene editing in the RANK/RANKL/OPG signaling pathway.
  
  

• Mechanism research on bone destruction in periodontitis.
  
  

• Construction of osteolytic models for cancer bone metastasis.
  
  

• Co-culture studies of immune cell-osteoclast interactions.

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