I am trying to grow a caco-2 cell monolayer on 96-well transwell insert plate (1 micron pore size) with a feeder tray at the bottom. The cell monolayer is grown for 21-23 days to allow tight junction formation. However, in the last week of my 21-day study, I observe a dark line in some wells in my plate as shown in the figure. I have also attached the photo of normal observed monolayer in the other wells in my plate. Is the dark line some kind of contamination or rupture of the monolayer or cell clumping/aggregation? I see no turbidity in the medium in the plate. I change the medium in the insert as well as the feeder tray every other day till day 14 and then almost everyday till day 21, because after day 14 the media changes color quickly due to tight junction formation. Media I use is Gibco high glucose DMEM supplemented with 10% FBS and 1% penicillin/sterptomycin antibiotic solution (10,000 units/ml penicillin and 10,000 ug/mL streptomycin). I don't observe such a dark line when I culture the cells in 24-well inserts.
Thank you.
Hi Eta,
This looks like classical dome formation in confluent Caco-2 cell line culture..
Good work
@Patrick: Thank you for your comments. However, I am a little skeptical that its a dome formation. I couldn't get the image of the entire well as I took the image at a higher magnification. I will try to post an image which shows the entire well. Its a single line which runs through the center of the well.
Ok Heta let's see.
In any case, I think very few domes are built in 96 well plates, wich could explain "single lines"..
I see. I wasn‘t aware that 96-well inserts show fewer domes. Thank you so much for your comments. I will post an entire image of the well next time, may be then we can be sure about it.
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