Hello Saeid.

Sorry! MSCs are hard to tell whether they differentiated into certain phenotypes unless you take further procedure with biomarkers of expression study. In particular, that magnification under phase contrast. This is probably due to the induced differentiation conditions are not perfect to drive all the cells to one direction of differentiation yet.

Regards.

Dear Saeid,

Some of them appear spindle shaped, like chondrocytes. Add human growth hormone (GH)and if they become swollen, they are chondrocytes. Anti-chondrontin sulphate strongly positive antibodystaining after GH also will do the trick,

Best,

AMB

When MSCs differentiate into condrocytes, they will form cartilage matrix and form micromasses, which stain blue with alcian blue pH 1. The pictures can be seen in this publication.

Jeanne Adiwinata Pawitan et al /Int.J.PharmTech Res.2014,6(1),pp 224-235.

Best way will be AB ph1 staining. Its difficult to say differentiation without staining.

You need to test gene expression using primers for collagen type 2, aggrecan, and perhaps Sox9. 2 weeks is a bit short, 3 is better. You can fix the cells and check proteoglycan deposition by either Alcuin blue or safranin O staining. again it’s better to do pellet cultures for this.

You should be doing micromass cultures. Chondrocye differentiation in standard 2D plastic adherent cultures is impossible. Look for Collagen 2 and Aggrecan.