Cytotoxicity test with simplified crystal violet staining method using microtitre plates and its application to injection drugs

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Abstract

A cytotoxicity test with the crystal violet staining method was developed using Chinese hamster lung and HeLa S3 cells in 96-well microtitre plates with an automatic plate reader, to facilitate examination of serial dilutions of the test substance. A good correlation was observed between this procedure and routine cell counting. However, for some chemicals results of the microtitre plate method differed from colony formation counts, particularly when the chemical induced cell stasis. In an application of the microtitre cytotoxicity assay to injection drugs, some drugs that are widely used proved to be very cytotoxic.

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    After incubation, the medium was removed, and the monolayer cells were washed with 100 μL of HBSS. Then the viability of the lung cell lines was measured by a colorimetric method as described in the literature (Itagaki, Hagino, Kato, Kobayashi, & Umeda, 1991; Saotome, Morita, & Umeda, 1989) in the presence of 1 % (w/v) of crystal violet dye which was added to each well for 30 min. The colorimetric intensity of sample was examined at 595 nm using a microplate reader, comparing with cell lines without the addition of tested sample as a control.

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