Title: Simultaneous determination of multiple end points in the screening of cytotoxic drugs, using laser-scanning fluorescence microplate cytometry.
Abstract: AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA
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Aim: To determine whether fluorescence microplate cytometry can provide multiple data end points for assessment of cytotoxic drug action on adherent tumour cell lines in vitro .
Methods: The effects of three different cytotoxic agents (5-fluorouracil, cisplatin and irinotecan) on six cell lines (MCF-7, HT-29, A549, A375, PANC1 and LNCAP) were assessed. Cells were grown in 96 well plates. When in log-phase growth, cells were transferred to reduced serum medium and treated with cytotoxics. In live cells, at set times after treatment, multiple fluorescence-based methods were employed to measure cell viability, apoptosis and cell cycle parameters in both unfixed and fixed cells. Data acquisition and analysis were performed using an Acumen Explorer fluorescence microplate cytometer (TTP Labtech Ltd., Melbourn, Herts, UK.)
Results: Data on nuclear size, nuclear DNA content, BrdU labelling, caspase activity and plasma membrane activity could all be obtained for each adherent cell within a culture well. Dose-response relationships for each cytotoxic agent were determined using these data. Data outputs could be obtained in the form of graphs, true-colour well images and false-colour heat map images. Table 1 shows representative data for relative cell number, relative nuclear size and relative nuclear DNA content obtained from HT-29 cells, 48 hours after treatment with 5-FU.
Conclusion: Microplate cytometry can provide data on cell number, cell viability, apoptosis and cell cycle simultaneously in a high throughput format. This has been demonstrated in a range of cell lines exposed to different cytotoxic agents. The methods described greatly facilitate the rapid screening of novel agents against a range of standard standard cytotoxic agents.
![Table][1] [1]: pending:yes
Publication Year: 2007
Publication Date: 2007-05-01
Language: en
Type: article
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