Cell culture using laminin - (Jan/20/2008 )
I'm very new to cell culture (ie this is my first one!) I am culturing mouse DRGs for calcium imaging.
The plates I am culturing on have a glass circle attached to a coverslip so that the culture can be used directly for the imaging experiment (which forms a kind of mini bath inside the normal culturing plate). The plates are glass and acid-etched and I used laminin in the centre of the glass circle and added my cell suspension. I then incubated for 2 hours before flooding the plate with 2ml of media.
After I had flooded the plate I thought maybe I'd made a mistake, because I flooded the entire plate (ie the mini bath and the culture dish) will this affect where my cells are or will they already be attached to where I applied laminin so will remain there even when I added more media?
Sorry if this is a bit confusing and I hope it makes sense and someone is able to help! I'm worried I've wasted a whole day (and a Sunday at that!)
Normally, we add laminin to coverslip and leave it overnight. The next day after a wash, we add cells. If you add the cells, and let them settle down, then flood the well, there might be a few cells, that might not stick but most of them should stick in the place of laminin. if the plates are not coated evenly with laminin or other chemicals, you can notice difference in density.