Label-free, real-time live-cell assays for spheroids: IncuCyte® bright-field analysis

A growing body of evidence suggests that more relevant and translational observations can be made with micro-tissues and organoids compared to 2D monolayer cell models. This is most notable in the cancer biology and hepatotoxicity field. For example, tumor spheroids exhibit more relevant morphology and increased cell survival compared to 2D cultures and have a hypoxic core. Current methods for assessing the growth and shrinkage of tumor spheroids are limited by one or more of the following: (1) assay workflows that are time-consuming, expensive or laborious, (2) a requirement to label the cells (e.g. a fluorescent probe) which may perturb the biology and may not be amenable to primary tissue, (3) single time-point readouts that do not report the full timecourse, (4) indirect readouts (e.g. ATP) that may overlook valuable morphological insight and/or mis-report cell growth. 

In this application note we describe methods and validation data for miniaturized (96/384-well) live-cell tumor spheroid assays that are based on non-invasive bright-field image analysis performed with the IncuCyte® S3 Spheroid software module. Tumor spheroids are formed in ultra-low attachment (ULA) plates and monitored for size and morphology for up to 2 weeks. These assays are flexible, simple to run and provide automated and direct measures of tumor size in real-time.