FACS Sorting and Protein Purification
Posted 19 September 2011 - 04:49 PM
I would like to ask if someone can pinpoint me reasons for decreased cell viability and moreover, how to avoid it. I would like to do some protein extractions from my sorts, but usually I have very low levels of alive cells and would like to try any suggestions.
Also, I am working with soft tissue, and I have read that for single cell isolation people recommend trypsin for ECM digestion although somebody told me its not a good idea to add it if I am going to do protein extraction. What do you think?
Posted 19 September 2011 - 08:35 PM
We use non-enzymatic celll dissociation solution to dislodge cells from adherent-monolayer cell-culture.
Posted 20 September 2011 - 02:26 AM
Posted 20 September 2011 - 04:37 AM
Posted 21 September 2011 - 05:57 PM
My nozzle size is 70um, although how changing the nozzle size would help to obtain more cells? They are already dead once you start doing the sorting, since I am staining with PI. Although if I am wrong, which could be for sure, please correct me!
Posted 28 November 2011 - 08:14 PM
Regarding the Trypsin thing. We use a product called Accutase, which is also pretty gentle on cells.