Preparing a Cell Suspension from the Adherent Skin Cell Cultures

To begin, remove the respective medium from the culture flasks containing the keratinocytes, fibroblasts, and melanocytes. Gently wash the cells with five to 10 milliliters of PBS. Now add 0.5 to two milliliters of Trypsin-EDTA solution to the flask, depending on its size.

Incubate the flask at 37 degrees Celsius. Use a microscope to control the detachment of cells from the surface. Suspend the detached cells in double the volume of Trypsin Neutralizer to deactivate Trypsin.

Then transfer the suspension into a 15-milliliter tube. Now pipette about 20 microliters of the skin cell suspension into a 1.5-milliliter tube. With the help of a hemocytometer, count the cells Next, centrifuge the tube at 300 g for five minutes at room temperature.

Then pipette out most of the supernatant and resuspend the cell pellet in the small amount of remaining liquid. Add an equal volume of fresh medium to the resuspended cells. In a separate 15-milliliter tube, prepare cell suspension.

The primary cells of keratinocytes, melanocytes, fibroblasts, and mast cells presented their typical morphology when grown in their respective media.