hi im working with podocytes. I am intending to perform IF on these cells.
But one puzzling thing happened. 6hrs post splitting the cells into the plate meant for immunoflorescence, i notice the cells have not adhere to the coverslips. I am using pre-coated poly-lysine coverslips. do cells generally take longer time to adhere to coverslips(pretreated or not pretreated) vs culture plate surfaces?
i also havent rule out possibility that i might have accidentally inverted the coverslip when splitting the cells just now. even with the non-coated coverslip side on, will cells eventually adhere to the slips?
Normally, there is no need of coated coverslips for cell adherence. The simple way to get rid off your problem is to keep cells on coverslips for longer time. Leave the cells overnight and check. Cells should be adhere very well. Good luck!
Cell adhesion to glass coverslips depends on cell types and coverslips pretreatment. Do you know if your cells like polysine?
Coverslips can have residue on them that prevent cells from adhering properly. We wipe ours clean with distilled water and a kimwipe. You can use ethanol to clean them if you need them to be sterile. Poly-L lysine is good for adhering cells non-specifically and should occur quickly if it will work. If you are plating your cells with serum, no coating is needed. If not, you may need to coat with extracellular matrix proteins such as fibronectin (10-15 ug/ml) or collagen (25ug/ml; just DO NOT use suction when rinsing the coverslips).
True, try to change coverslip from other company, some charge may vary. Try poly-lysine coated also. Giving more time will help as Subrata mentioned- That is important too
Hi,
What is the suitable coverslips coating reagent for HCT116 cell line?
Thank You
I use autoclaved normal coverslips, confocal dish (VWR) and Lab-Tek chamber slides (Thermofisher) for both immortalized human and mouse podocytes. The plating container bottom is pretreated with rat tail collagen I in 37℃ incubator for 1 hour. Trypinizing of differentiated cells is shorter than undifferentiated cells, or they may not adhere. Usually our podocytes adhere in 1 hour and adhere very well over night.
Coating solution:
0.02N Acetic Acid (Sterile): 49 ml
Rat tail collagen I: 1 ml (BD Boscience, #354236)