All in one microplate reader

Total RNA was extracted from BSC cultures using the Absolutely RNA Microprep Kit (Agilent Technologies, Cedar Creek, TX, USA) as per the manufacturer’s protocol. Cells were lysed at 0, 2, 4, 8, 12, 24, and 48 h post-treatment. In brief, lysis buffer was added directly to the culture dish and a cell scraper was used to further lyse the cells. The cell lysate was then vortexed and an equal volume of 70% ethanol was added. The mixture was then centrifuged and filtered. A series of wash buffers were added and then the RNA was eluted and stored at −80 °C. Isolated RNA was quantified using a BioTek all-in-one microplate reader using Gen5 2.0 software (BioTek Instruments, Winooski, VT, USA), and quality was determined using the 260/280 ratio. Samples with a ratio greater than 2.0 were deemed high enough quality for cDNA synthesis. All RNA samples were treated with deoxyribonuclease (Ambion, Foster City, CA, USA) before beginning cDNA synthesis using a high-capacity cDNA reverse transcription kit (Applied Biosystems, Foster City, CA, USA) following the manufacturer’s protocol.