In Vitro Cytotoxicity Assays and IC-50 for 120 Tumor Cell Lines
Altogen Labs offers a number of cell-based tissue culture assays, such as flow cytomerty and microplate-based cell proliferation and cytotoxicity assays (including cell cycle, cell viability, membrane protein expression, TUNEL, caspase 3/7 apoptosis, etc). Standard MTT cytotoxicity assays are based on the cleavage of the yellow terazolium salt (XTT) by metabolically active human cells (usually HeLa or A549 cells) to form an orange formazan dye. This conversion only occurs in viable cells, so cytotoxicity leads to a loss of activity. The formazan dye formed is soluble in aqueous solutions and is directly quantified using a scanning multi-well plate reader (spectrophotometer). Multiple replicates ensure that this method provides a high degree of accuracy and statistical data confidence. The assay is suitable for the performance of dose-response studies and is offered in a 96-well microplate format.
Cytotoxicity assays can be necessary for a variety of reasons; initial drug screenings require a thorough understanding of how a given drug impacts cells, and other kinds of testing may require knowledge of cellular responses to introduced reagents. Cytotoxicity assays provide the information necessary for determining future screenings of drugs in vivo, and can also give valuable insight into the cytotoxic effects of other compounds. Such information can inform pre-clinical trials, and will also give an early indication of poisonous properties of compounds.
IC-50 testing service is offered in over 120 in-house cell lines and primary cell types or any custom (client provided) cell line. The goal of this test is to measure the biological activity of a compound of interest (IC-50) against different types of tumor cell lines. In standard setup, cells in logarithmic growth are seeded in 96-well tissue culture plates (5,000-7,500 cells/well). Appropriate negative and positive controls are used in the assay (control conditions usually include the nontreated cells, cells with medium and vehicle as a negative control and one of the standard chemotherapy drugs as a positive control). Plates are assayed at 48-72 hr after initiation of compound exposure, then a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) solution is added to each well for a 1- to 3-h incubation (at 37°C and 5% CO2). Analysis is performed at least in triplicate (biological replicate) and using a plate reader to generate IC-50 assay data.
The IC-50 assay is incredibly important in cancer research, as it provides quantitative information regarding specific dosages of drugs and their effects on cell lines. For cancer drug screenings, the IC-50 assay is an absolute necessity, as it can be used to determine the concentration of a drug to be used for in vitro screenings, which will precede clinical trials. The assay requires large amounts of individual wells to be used for different concentrations of drugs, and as such it requires considerable precision and effort to complete successfully. As such, reproducibility can be greatly increased if the assay is conducted by a company with pre-determined standard procedures for its execution.
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