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Length of incubation for transwell migration assay


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#1 microRNAboy

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Posted 22 November 2010 - 04:01 AM

Hi All,

I've been having some difficulty with my migration assays, using BD transwell inserts.

I was previously using Corning plates (but have switched due to price) and found that a 4-hour incubation was fine to be able to measure migration of cancer cell lines towards 20% serum.

I've been trying to optimise my incubation time and have found that 5 hours is too short (many cells are 'stuck' in the membrane pores and not many are fully through) and that overnight incubation is far too long (the cells form a monolayer on the underside of the membrane).

I realise that I will have to use an incubation time between these two points, but I was wondering what other peoples experience has been with incubation length. It seems from the literature that 4 hours is fairly standard.

Thanks!

#2 ElHo

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Posted 23 November 2010 - 04:20 AM

Incubation time depends on the migratory potential of the cell line used and on the chemoattractant. So I donīt think that there can be a standard value and you have to optimize it for your conditions. We usually incubate 22h using a highly metastatic breast cancer cell line.

#3 microRNAboy

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Posted 23 November 2010 - 04:55 AM

Incubation time depends on the migratory potential of the cell line used and on the chemoattractant. So I donīt think that there can be a standard value and you have to optimize it for your conditions. We usually incubate 22h using a highly metastatic breast cancer cell line.


And is that for migration or invasion? If you don't mind me asking, what do you use as chemoattractant?




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