Which protocol should I use for dissociation of hepatocytes?

iCell® Hepatocytes 2.0 are amenable to enzymatic dissociation of the monolayer from a culture vessel for subsequent replating into an assay-specific plate. Gentle dissociation is required to preserve health and platability of the cells. We have optimized the dissociation for iCell Hepatocytes 2.0 using TrypLE Express Enzyme (Thermo Fisher Scientific, Cat. No. 12563011) or StemPro Accutase Cell Dissociation Reagent (Thermo Fisher Scientific, Cat. No. A11105). Allow digestion to continue at room temperature until cells just begin to detach from the plate (approximately 2 - 4 minutes). See our Application Protocol for more details.

Although we generally do not recommend using a trypsin dissociation reagent because it is harsher on cells and leads to more cell death, we have tested 0.1% trypsin/EDTA for 3 minutes, which resulted in singularized cells. Therefore, if singularized cells are a priority for the application, we recommend you optimize the trypsinization conditions with the understanding this reagent can limit cell re-attachment in 2D and 3D applications.

Reference:

  1. Application Protocol: Dissociating Hepatocytes by Enzymatic Treatment