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transwell inserts


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#1 biologg

biologg

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Posted 12 July 2011 - 10:02 AM

Hello everyone,

I would like to ask you for an advice. Let me first explain exactly what I am doing: I want to perform a migration assay using Transwell inserts from Corning, looking at muscle myoblasts (an adherent cell line). I have resuspended my cells in serum-free media and allowed them to migrate for 4h towards serum-containing media in the receiver plate or serum-free media (as a negative control). In a situation when the inserts have not been coated with an ECM, as expected, there are almost no cells on the bottom part of the membrane, when serum-free is present in the receiver plate, and many cells when serum-containing media is present in the receiver plate. This is obviously looking at the chemotactic response of cells towards different chemoattractans present in the receiver plate. However, when i coat the transwells by diping them in 0.2% gelatin for 30min at RT, 4h later there is a huge amount of cells migrated in the well with serum-free present in the receiver plate - in fact, much more cells compared to the well with serum-containing media (these results are reproducible). Anyone knows what is going on? My explanation would be that the cells would migrate towards the ECM on the bottom part of the membrane, even if no chemoattractant is present, but i cannot explain why there are more cells in the receiver with the serum-free media compared to the well with serum-containing media. Can anyone tell me why this is happening?




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