9
Gelatin Coating
Preparation of gelatin solution:
1. Add gelatin powder to PBS at a concentration of 2%
2. Autoclave the mixture to dissolve and sterilize
Coating with gelatin solution:
1. Pour 3 ml of the gelatin solution into each strain chamber
2.
Incubate at 37
o
C for more than 30 minutes
3. Aspirate the gelatin solution. If coating is successful, water will not be repelled after removing
the
gelatin solution.
4. The liquid solution can be used to coat 3 or 4 chambers before discarding.
Collagen Coating (Cellmatrix 1-C, P, Type 3 or 4)
Preparation of collagen solution:
1.
Combine 1 part collagen to 10 parts HCL, pH 3, in a sterile
tube
Coating with collagen solution:
1.
Coat chamber with a thin layer
2.
Aspirate excess
3.
Dry in biological safety cabinet at 25°C or below. The
chamber can be stored at the same temperature.
4.
Wash the chamber twice with culture medium.
5.
If coating is successful, water will not be repelled.
The PDMS (silicone) chambers are
very hydrophobic with two
methyl-bases on the surface. Cells
adhere to the fibronectin or collagen
coated chambers via integrins. This
form of cell adhesion is very different
compared to attachment of cells to
plastic or glass dishes where the
surface of plastic or glass is charged,
resulting in non-specific binding.
If the cells are having difficulty
attaching to the coated chamber or
easily detached upon stretching, treat
the stretch chamber with a higher
concentration of fibronectin or
collagen, for overnight.
