I am currently and undergraduate taking tissue culture at my university. We are working with HEK293 cells and thus far have not had any problems with contamination or with my 10% FBS + PSG MEM media. We started using the really small tissue culture plates with 1ml of media with my cells and placed them in 37 incubator. I’m plotting a growth curve of my cells over the course of a week but half my plates look really good and half look balled up and not adhering to plate. My day 1, 2 and 3 cells looked great but my day 4 and 5 plates, my media dried up and looked crusty?? There wasn’t any mold on the plate and the incubation temperature and co2 looked fine. I’m just curious as to why half my plates look fine and half have dried up.
Hi Kelsey
What type of culture plate are you using? Is it a 6-well, 12-well, 48-well or 96-well plate?
I have also experienced this drying issue before in a 37 degree incubator. The trick is to surround the outermost border wells (e.g. first row; last row, first column, last column) with maximum working volume of PBS/dH2O. By doing that, these buffering solution will evaporate first instead of your media containing cells.
Hope it helps and good luck!
I think its pretty odd that your media has completly dried up after 4 days...make sure you are using the correct volume of media for the type of plate you are using. Also, make sure there is water in the water pan that should be probably at the bottom of the incubator...lastly you can do as Shou Jun Phang suggests.
Hello Kelsey Salazar
During culture, media commonly evaporate from the wells that are closest to the perimeter of the plate, with the outer and corner wells being the most affected. This result in variation in cell growth across the plate. Besides what has been mentioned by Shou Jin Phang it is important that culture plates are incubated in an environment with at least 95% humidity. Evaporation is almost four times higher at 80% than 90% humidity, demonstrating the significant difference that humidity levels can make on resulting viability of cells.
Another critical factor affecting evaporation is the frequency with which plates are removed from the incubator for microscopic examination. Furthermore, as is often the case, incubators can be shared by multiple users, making frequent door openings another issue. Exposure to external conditions decreases humidity control and temperature within the incubator, thus increasing the rate of evaporation. Please bear these points in mind whenever you work in cell culture.
Best.
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