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tranwells and C6 glioblastoma cell migration - (Jun/05/2008 )

Hello everybody!

I am doing for the first time a migration assay using C6 cell line and 8 um-transwells. From what I had read on this type of assay I was expecting cells to remain attached to the lower side of the membrane after migrating through it. Instead, I found lots of cells attached to the bottom of the well (usong 6-well plates). So my questions are:
1) is this "normal"? Should I just count cells on the bottom (burker chamber or cellular staining, which better?!) and would that be sufficient to quantitate migration?
2) could this phenomenon be caused by bad choice of pore size? C6 cells are rather small
3) could it be something wrong with use of the transwells? I followed instructions and wetted the membranes on both sides.

Thanks a lot to anyone who can give me any help & advice!

-mascat-

QUOTE (mascat @ Jun 5 2008, 07:09 AM)
Hello everybody!

I am doing for the first time a migration assay using C6 cell line and 8 um-transwells. From what I had read on this type of assay I was expecting cells to remain attached to the lower side of the membrane after migrating through it. Instead, I found lots of cells attached to the bottom of the well (usong 6-well plates). So my questions are:
1) is this "normal"? Should I just count cells on the bottom (burker chamber or cellular staining, which better?!) and would that be sufficient to quantitate migration?
2) could this phenomenon be caused by bad choice of pore size? C6 cells are rather small
3) could it be something wrong with use of the transwells? I followed instructions and wetted the membranes on both sides.

Thanks a lot to anyone who can give me any help & advice!


think of a good coat for the lower chamber to attract cells; for smaller cells 5 or even 3 ┬Ám pore size may be enough

-The Bearer-