I am constantly getting variation in the data set. Need some suggestions if anybody can share their experience.
This edge effect is likely the result of differences in evaporation and thermal changes in the plate.
One of the easiest tricks, is to put blank media or sterile water in the edge wells. In my experience, this works well, but of course wastes some of the plate, which can be inconvenient. Another solution is to use a gas permeable sealing film, to reduce evaporation.
Recently, I saw a post here linking the following paper:
http://www.ncbi.nlm.nih.gov/pubmed/14567784
They show nice results eliminating edge effete by letting the cells adhere for an hour at room temperature before placing the plate in the incubator. The adhesion process is then isothermal, so variability should be reduced. I personally have not had a chance to try this, but it seems like a simple thing to give a whirl with your cells and see how it goes.
After seeding the plate, I incubate it in laminar hood for more than one hour. once the cell are attached to the substratum then only i transfer them to the incubator. This i have read in one of the article from a reputed journal but i think its not working in me.
This edge effect is likely the result of differences in evaporation and thermal changes in the plate.
One of the easiest tricks, is to put blank media or sterile water in the edge wells. In my experience, this works well, but of course wastes some of the plate, which can be inconvenient. Another solution is to use a gas permeable sealing film, to reduce evaporation.
Recently, I saw a post here linking the following paper:
http://www.ncbi.nlm.nih.gov/pubmed/14567784
They show nice results eliminating edge effete by letting the cells adhere for an hour at room temperature before placing the plate in the incubator. The adhesion process is then isothermal, so variability should be reduced. I personally have not had a chance to try this, but it seems like a simple thing to give a whirl with your cells and see how it goes.
I think this will help you:
http://www.labnews.co.uk/features/beating-the-edge-effect/
I don't use the perimeter wells for the experiment, and I fill them with either medium or extra cell suspension. Also make sure your incubator is well humidified to minimize evaporation.
I had such issues with MTT due to evaporation. I used to test with just my media or cells in the last columns. That helped a lot to monitor any discrepancies if present. Am curious: Why do you incubate in the cabinet for a hr instead of placing them in incubator? and how long are u test periods?
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