Collagen coating of glass bottom dishes and glass bottom plates

Materials

Rat tail collagen (type I) solution

Rat tail collagen I is available from many commercial vendors, such as BD Biosciences(cat# 354249), Sigma (cat #C3867), Invitrogen (cat #A10483-01).

It is recommended to purchase collagen solutions, so that sterilization is not needed since filter sterilization of collagen could be problematic.

Dilution the stock solution to 50 ug/ml with 0.02M acetic acid or 0.01M HCl.

Procedure

  • Add solution (50 ug/ml) into a glass bottom dish or glass bottom plate at 5 ug/cm2. If cells will be cultured only on the glass surface, then only the glass surface needs to be covered with collagen solution.
  • Incubate for 1 hour at room temperature in a hood.
  • Carefully aspirate the collagen solution.
    (alternatively, the glass bottom dish/plate can be left in the hood to dry overnight, and the following washing step is not needed)
  • Rinse 3 times with PBS or serum-free media.
  • Collagen coated glass bottom dish or glass bottom plate can be air dried and kept at 4℃ for about a month. Note that coating on glass surface is less stable than coating on plastic tissue culture dishes or plates. Thus it is recommended to coat the glass bottom dish/plate right before cell culture.

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