Cell viability was assayed using either the CellTiter AQueous Non-Radioactive Cell Proliferation Assay (MTS, Huntsville, AL, USA) or CellTiter Luminescent Cell Viability Assay (Promega, Madison, WI, USA) as described previously.18 (link) BrdU incorporation was assayed using a BrdU Cell Proliferation Assay according to the manufacturer's suggested protocol (Calbiochem, Billerica, MA, USA). Real time analysis of cellular proliferation by xCELLligence assay was performed as described previously.18 (link)Apoptosis was determined by nuclear fragmentation assay as described previously.54 (link), 55 (link), 56 (link), 57 (link) Quantifications were performed in triplicate, with each count consisting of at least 300 cells. Caspase 3/7 activation was evaluated by Caspase 3/7 kit (Promega) according to the manufacturer's suggested protocol.
Caspase 3 7 kit
Manufactured by Promega
The Caspase 3/7 kit is a laboratory assay designed to measure the activity of caspase-3 and caspase-7, two important enzymes involved in the apoptosis (programmed cell death) pathway. The kit provides the necessary reagents and protocols to quantify the cleavage of a luminogenic substrate specific for these caspases, allowing researchers to assess cellular responses and apoptotic processes.
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Lab products found in correlation
2 protocols using caspase 3 7 kit
1
Cell Viability, Proliferation, and Apoptosis Assays
2
Caspase-3/7 Activity Assay for Apoptosis
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Caspase Glo® Assay kit (Promega) was utilized in order to measure apoptosis by proxy of caspase 3 and 7 activity. This was carried out in order to further characterize the mechanism of DNMTi drug induced apoptosis. Cells were cultured in the above-mentioned conditions, and the luminescent signal was assessed using the manufacturer’s instructions. The readout for each drug concentration was determined as a relative percentage to the control ((experimental/control) *100) and plotted using Graph Pad Prism.
Furthermore, levels of apoptosis for the guadecitabine treated LMS cells were quantified during the delayed response experiment using the abovementioned Promega Caspase 3/7 kit. Following treatment, cell lines were reseeded in drug free media in 96 well plates. The plates were incubated in the same conditions as the MTT delayed response experiment for 1, 2, or 3 days. Luminescence was measured as previously described above.
Furthermore, levels of apoptosis for the guadecitabine treated LMS cells were quantified during the delayed response experiment using the abovementioned Promega Caspase 3/7 kit. Following treatment, cell lines were reseeded in drug free media in 96 well plates. The plates were incubated in the same conditions as the MTT delayed response experiment for 1, 2, or 3 days. Luminescence was measured as previously described above.
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