Page 93 - Live-cellanalysis handbook

Kinetic Single Spheroid Assays

®
Application of IncuCyte S3 Single Spheroid live-cell assays
IncuCyte® S3 DF Brightfield Image Acquisition (described in Brightfield signature is obtained which, when combined with
IncuCyte S3 Multi-Spheroid Assays, Figure 2) enables robust fluorescence analysis using IncuCyte® Cell Health Reagents,
measurement of spheroid size, yielding information on spheroid provides insight into the viability of tumor spheroids in response to
growth rates and morphology without the need for labels. different treatments.
Following treatment with cytotoxic compounds, a strong

Label-free monitoring of spheroid growth and morphology over time

The example below illustrates how the IncuCyte S3 DF Brightfield object in the field of view. Changes in the size of spheroids derived
analysis can provide kinetic measurements of single spheroid size from three types of tumor cell were monitored in response to
over time (Figure 2). The size of tumor spheroids was measured camptothecin, allowing growth response curves to be generated in
using an automated algorithm that masked the largest Brightfield the absence and presence of the drug.

Breast Connective Tissues Lung
MDA-MB-231 + Matrigel HT-1080 A549

Vehicle

CMP (1µM)

Figure 2. Brightfield analysis enables accurate kinetic quantification of spheroids. The differential pharmacological effect of 1 μM camptothecin (CMP) on
growth of MDA-MB-231, HT-1080 and A549 cells in a 3D spheroid assay. Cells were grown in ULA round-bottom 96- well plates (2,500 cells per well) for 72
h and treatment with ± 1 μM CMP followed. Segmented DF Brightfield images compare treated vs. un-treated conditions at 240 h. Time courses illustrate the
2
specific cell type-dependent kinetic profile of spheroid growth and shrinkage. The graphs display the Largest Brightfield Object area (μm ) (y-axis) over the
course of a 240 h assay (x-axis) at 6 h intervals. All images captured at 10x magnification. Each data point represents mean ±SEM, n=4.

91