Help with improving scratch migration assay protocol - (Jan/17/2007 )

Hi all

I'm doing a scratch migration assay, and one thing I am having trouble with is making a clear, definite scratch with defined borders -- I am using a pipette right now but the scratch doesn't seem to be clean enough.

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depends also on the type of cells; if they strongly kohere, it is difficult to get a clean line

an alternative migration assay is using cell culture plates with filter pores and a second lower chamber to trap migrated cells from the upper chamber

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Thanks Bearer. The transwell migration assay is actually my next experiment, but I have to do both. I am using Rat-2 cells. Any tips in general???

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Hi

I'm doing this assay too.... I'm not sure till now if it is better to do it in serum free medium or low serum,,, which one do u think is better. what is the best way to quantitate the results....

thanks

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I use a tip (1 ml) to make the scratch. It works fine to me. Another guy in the lab uses a sterile razor, with help of a little rule (of course sterile too). Lean the rule on the side where cells will be deattached and do 2 scratches separated by some few milimeters just to have a wide scratch of width desired. Finally deattach the cells that could remain attached in the middle of the scratch with a tip. The scratch if very well difined but the procedure is a little hard to me. Maybe it's matter of practice. Moreover the cells I'm using are bigger than the other guy's, so he need a more defined scratch.

About the media, it's depends on the cell type. Some cells need at least 0.2% serum in media to achieve some migration. other are ok with no serum. Do a couple of tests to check it.

Good luck!

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