I have different test groups, testing for MG-63 and the viability and proliferation tested by MTS assay.
samples revealed the cell viability ( (ODsample/ODTCP)*100 ) lower than 70% by 2 days. However, since I am doing the cell test for 7 days, the cell viability increased higher than 70% after 4 days. can I say the samples were not cytotoxic?
I'm afraid you can't say that. The samples might be killing 20% of the population that gets replaced in 4 days, without a change in OD. When cells are confluent they're more resistant to stress. So it's a good idea to test your cells at different plating efficiencies.
Cytotoxicity is possible without cell death. Genotoxic compounds with carcinogenic potential would be a good example.
Oscar L. Sierra Thank you for answering. So you would say that the samples are cytotoxic and the cell viability higher than 70% after 3-4 days didn't mean anything?
If I'm not mistaken, MTT and other assays are surrogate measures of normal mitochondrial enzyme activity. The assay you're running gives you a denominator for the number of cells with normal mitochondria. You could measure other markers (stress markers, changes in heat shock proteins, oxidative stress, etc) in the numerator and correct for number of viable cells. Each organelle might have its own stress markers (for example for protein aggregation that might be cytotoxic might trigger the ER stress pathway, autophagy and ERAD-proteasome pathway. For genotoxic aggents there are assays for chromosome instability, DNA degradation, up-regulation of DNA repair machinery, etc. For teratogenic agents, you need an animal model. For apoptosis there are other markers like caspase 3 assays, TUNEL assays, transglutaminase activity. For telomere shortening there are corresponding assays, and so on. - Look for the type of agents present in the sample and test the most likely cytotoxicity pathways. If cells are grown at low plating efficiency but not so low (can be detrimental for cell growth), they adapt for cell cycle stage S, and so their mitotic rate could be relevant to test (cell arrest decreases upon toxicity in S phase or mitosis). I hope it helps
Cell Line Description
From an osteosarcoma of a 14 year old male. Produces high yields of interferon following superinduction with polyinosinic acid-polycytidilic acid, cyclohexamide and actinomycin. Antigenically, MG-63 interferon is more closely related to human fibroblast than to leukocyte interferon.
Application
Human MG-63 cell line has been used:
• to investigate the cytotoxicity of acrylic bone cement extracts on MG-63 cells
• to fabricate three-dimensional (3-D) niches and study the cell viability, adhesion and proliferation of MG-63 cells
• as a positive control for immunostaining procedure
• to study the interactions and effects of lysophosphatidate functionalized titanium on MG-63 cells
Interferon-induction studies
-- Subculture Routine
Split sub-confluent cultures (70-80%) 1:3 to 1:6 i.e. seeding at 2-4x10,000 cells/cm2 using 0.25% trypsin or trypsin/EDTA; 5% CO2; 37°C.
--NOTE
cyclohexamide (inhibits protein synthesis) and actinomycin D (inhibits RNA synthesis) are cytotoxic drugs.
Oscar L. Sierra Yes MTS assay is to assess the cell proliferation, which the quantity of formazan product as measured by the amount of 490nm absorbance is directly proportional to the number of living cells in culture. So I though it can be an indicator of cytoxicity as well
Oscar L. Sierra TCP was the control, and the samples were in the ultra-low attachment well plates, to minimize the attachment of the cells to the wells.
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