When should I use ROCK inhibitor?

A common question we get is about when should media contain ROCK inhibitor (molecule Y-27632)?

As a rule of thumb, cells should be fed media containing ROCK inhibitor 24 hours after thawing, and for 24 hours after each single cell passaging event.

ROCK inhibitor can also be used for a few hours before a “traumatic” cell event like FACS enrichment or transfection to boost viability.

Have you tried not using ROCK inhibitor if you passage as colonies instead of single cells?

Hello Adriana!

I’ve included an answer to this question in another thread of yours, but for folks just joining this one, I’ll answer here as well. While I have not tried this personally, my teammate Angel is very familiar with clump passaging and said the following:

While passaging in clumps, you do NOT need ROCK inhibitor. At a freeze or a thaw step, you can use it for added survival, but not at the regular passage steps. One thing to add: if you’re using dispase or collagenase to dissociate, you should add in an extra wash with PBS or media before plating cells.

I hope this helps!

See related thread: Single cell vs colony passaging