Hello,
I work with 21-day differentiated Caco-2 cells and I want to do a trypan blue count.
Unlike undifferentiated cells, trypsin (i use TrypLE Express to be exact) does not allow me to separate cells from each other and I get very large cell clusters.
What could I use to separate them effectively without inducing rapid cell death?
Thanks in advance
Dear kevin
My rwcommendations
1. Be sure you wash your cells with PBS Ca/Mg free to fragilize the cadherins.
2. Use fresh trypsin/EDTA 0.25%.
3. Increase your incubation time in trypsin until you have good cell separation.
4. Keep your cell passaging less than 70% confluency otherwise you want them to form monolayers and have them undergo differentiation.
In addition, when resuspending you might want to do it in complete medium. And I usually pass them 4-5 times through syringe (22 gauge).