Hello Chen wei Chen

Yes, you may have to modify the method.

DMEM + 10% FBS + 1% PS may not be sufficient for the development of chondrocyte. Though FBS contains the essential extracellular matrix proteins for cellular adhesion and provides relevant hormones, enzymes, and growth factors for cellular proliferation, it has been reported to affect chondrocyte biosynthesis in monolayer culture. Studies have shown that FBS would result in excessive cell proliferation or chondrocyte phenotypic instability.

Therefore, you may supplement the culture media with commercially available Insulin, Transferrin, and Selenium containing premix solution (ITS) consisting of 5mg/ml Insulin, 5mg/ml Transferrin, and 5μg/ml Selenious acid. Add 1ml of this solution for 1 litre of media. ITS will stimulate cell proliferation while decreasing substantially the serum requirements.

Instead of 10% FBS, you may use small volume of FBS along with ITS. To choose an appropriate FBS concentration, culture media may be supplemented with a combination of different concentrations of FBS and ITS. For instance, you may try the following combinations and choose the right combination that provide optimal results.

1% ITS plus 2% FBS,

1% ITS plus 4% FBS,

1% ITS plus 8% FBS, or

medium plus 10% FBS.

The below attached paper will be helpful to design your experiment.

Article Optimum Combination of Insulin-Transferrin-Selenium and Feta...

It is also preferable to add L-glutamine to complete culture media. This essential amino acid serves as a carbon source. For chondrocyte primary culture, you may add 1ml L-glutamine (from 200 mM stock solution) to 100 ml DMEM media. Add 1% PS to prevent contamination.

Good Luck!

Hi Malcolm Nobre

Thanks for your comment ! I will give it a try.