vitro cytotoxic study Biological activity for anti-cancer activity is created using the screening

600 WordsApr 23, 20193 Pages
vitro cytotoxic study Biological activity for anti-cancer activity is created using the screening process to guide the separation of crude extracts. The in vitro tests are usually used as primary screening in cytotoxic assay. 3-(4,5-dimethylthaizol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT assay) The MTT assay is based on metabolic reduction of colorless tetrazolium salt (MTT), by mitochondrial enzyme activity in viable cells, to formosan salt (blue), which can be quantified spectrophotometrically. It is particularly useful for assaying cell suspensions because of its specificity for living cells [8]. This assay was performed in 96 well plates at a cell density of 3x103 cells/well. Cancer cells precultured in media depend on type of…show more content…
The protein-bound dye is then solubilized by weak base for spectrophotometry. This colorimetric assay can be used to estimate the cell number indirectly by providing a sensitive index of total cellular protein content which is nearly related to cell density [12]. This method using cell line and described previously. After cell were incubated for 72 h after addition of test material. After incubation cells were fixed by gently layering trichloroacetic acid (50 µl/well, 50% w/v) on top of the medium in all the wells and incubated at 4 ºC for 1 h. The plates were washed and air dried. Cell growth was measured by staining with sulforhodamine B dye (0.4% w/v in 1% acetic acid, 100 µg/well). The adsorbed dye was dissolved in Tris-Buffer. The optical density (OD) was recorded using 96 well plate reader at 540 nm. Growth inhibition = 100 – [(OD test sample – OD blank / OD control – OD blank) x 100] [12] . Comet assay The single cell gel electrophoresis (comet) assay is an attractive test for measuring gene toxicity, investigating DNA repair, or monitoring population for exposure against environmental mutagens. The comet assay is widespread applications in gene toxicity testing in vitro and in vivo, human bio monitoring or eco toxicology [13]. Samples can be
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