Microplate spectrophotometry for high-throughput screening of cytotoxic molecules.
OBJECTIVE
High-throughput chemical and biochemical technologies are now exploited by modern pharmacology and toxicology to synthesize a multitude of new molecules with bioactive potential, or to isolate them from living matter. Testing molecules in cell systems on a large scale, however, is a rate-limiting step in drug discovery or in toxicity assessment. In this study, we developed a low-cost high-throughput method for first-level screening of cytotoxic molecules.


METHODS
We used microplate spectrophotometry to measure growth kinetics of human tumour cells that grow in suspension (Molt3) or adherent to the plastic surface of culture wells (HeLa) in standard RPMI medium. Cells were treated with colchicin, idarubicin or paclitaxel under various treatment schedules. The effects were quantified and compared with those measured by optical microscopy using the trypan blue dye exclusion method to reveal dead cells.


RESULTS
Proliferation kinetics of tumour cells can be quantified by measuring variations in optical densities of cell samples at 410 and 560 nm wavelengths. For cells that grow in suspension, one single reading at 730 nm may be sufficient to reconstruct growth curves that parallel those obtained by direct cell counting. Effects of the cytotoxic treatments could also be quantified and results compared very favourably with those obtained using standard techniques.


CONCLUSIONS
Microplate spectrophotometry is a robust and sensitive method to monitor growth of animal cell populations both in the absence and in the presence of cytotoxic drugs. This method implements existing technologies and can be fully automated.
Tomelleri
C
C
Department of Clinical and Experimental Medicine, University of Verona, Verona, Italy.
Dalla Pellegrina
C
C
Chignola
R
R
D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects*; D002448:Cell Adhesion; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003078:Colchicine / Q000633:toxicity; D003470:Culture Media; D003603:Cytotoxins / Q000633:toxicity*; D004847:Epithelial Cells / Q000187:drug effects*; D006367:HeLa Cells; D057166:High-Throughput Screening Assays / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D015255:Idarubicin / Q000633:toxicity; D007700:Kinetics; D015456:Leukemia, Biphenotypic, Acute / Q000473:pathology; D017239:Paclitaxel / Q000633:toxicity; D012680:Sensitivity and Specificity; D013053:Spectrophotometry / Q000295:instrumentation; D013601:T-Lymphocytes / Q000187:drug effects*; D013997:Time Factors
D003470:Culture Media; D003603:Cytotoxins; D017239:Paclitaxel; D003078:Colchicine; D015255:Idarubicin
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