Hi all,
We are currently seeding cells into alginate hydrogels, we are having the following issues, if anyone can help it would be greatly appreaciated!
1). Have any of you tried using dialysis tubing to form alginate plugs?
2). What is the best way to get even distribution of your cells, we try just pipetting up and down, and they look homogenous, but after we form our plugs and look under the microscope, the cells seem to stay more on the edges and not in the middle
2). After our experiments we need to isolate our cells for RNA extraction and Protein extraction. We have tried dissolving the alginate in both Sodium Citrate and EDTA. There are two issues: We need to dissolve them quickly because we are going to look at gene expression after a particular treatment (and it might change if it takes a long time to dissolve). Second, we seem to be losing a lot of cells somewhere. -- ie. if we seed say 5 X 10^ 6 cells/ml, make the plugs, then dissolve, we are only recovering a fraction of those cells. A lot of them are either completely lysed or dead.
3). For protein extraction, we are having issues with contaminating protein from the alginate, even after washing cells 2x with PBS.
Thanks!
Slkummer
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