title: Paired image- and FACS-based toxicity assays for high content screening of spheroid-type tumor cell cultures.
abstract: Novel spheroid-type tumor cell cultures directly isolated from patients' tumors preserve tumor characteristics better than traditionally grown cell lines. However, such cultures are not generally used for high-throughput toxicity drug screens. In addition, the assays that are commonly used to assess drug-induced toxicity in such screens usually measure a proxy for cell viability such as mitochondrial activity or ATP-content per culture well, rather than actual cell death. This generates considerable assay-dependent differences in the measured toxicity values. To address this problem we developed a robust method that documents drug-induced toxicity on a per-cell, rather than on a per-well basis. The method involves automated drug dispensing followed by paired image- and FACS-based analysis of cell death and cell cycle changes. We show that the two methods generate toxicity data in 96-well format which are highly concordant. By contrast, the concordance of these methods with frequently used well-based assays was generally poor. The reported method can be implemented on standard automated microscopes and provides a low-cost approach for accurate and reproducible high-throughput toxicity screens in spheroid type cell cultures. Furthermore, the high versatility of both the imaging and FACS platforms allows straightforward adaptation of the high-throughput experimental setup to include fluorescence-based measurement of additional cell biological parameters.
authors: Department of Surgery,, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors: Kari
authors: K
authors: Trumpi
authors: Cell Screening Core, Department of Cell Biology, Centre for Molecular Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors: David A
authors: DA
authors: Egan
authors: Department of Surgery,, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors: Thomas T
authors: TT
authors: Vellinga
authors: Department of Surgery,, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors: Inne H M
authors: IH
authors: Borel Rinkes
authors: Department of Surgery,, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors: Onno
authors: O
authors: Kranenburg
keywords: High-throughput; Spheroid-type tumor cell culture; Toxicity assay