Measurement of Cartilage Synthesis

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Chondrocytes will synthesize cartilage in vitro. This protocol describes how to measure and modulate it.

Materials

  • Bovine Chondrocytes (catalog # 1020 or 1021)
  • X VIVO 15® without Phenol Red (Lonza, Walkersville, MD)
  • L-ascorbic Acid (add to medium at final concentration of 50 ug/mL)
  • TGFß or Other Agent to Stimulate Production, If Desired (Phenol red cannot be used if the supernatants are tested using the DMB method. See sample data on the chondrocytes page.)

Procedure

  1. Thaw chondrocytes following recommended protocol.
  2. Adjust cell concentration to 1 million per mL.
  3. Add 100 ul to each well of a 96 well plate.
  4. Prepare agent to modulate cartilage synthesis in medium at twice the desired final concentration.
  5. Add agent to wells at 100 ul per well.
  6. Incubate at 37°C for 2–5 days.
  7. Remove supernatant and test for type II collagen or proteoglycans (see our proteoglycan detection kit).