Biotek synergy 4 plate reader

Cell proliferation was measured by Cell Titer-Glo luminescent cell viability assay (Promega, Madison, WI) according to the manufacturer’s instructions. 2 × 103 bladder cancer cells/well were cultured in 96-well white plates in media for 24 hours, then treated by compounds for 72 hours. Luminescent signals were read on the BioTek Synergy 4 plate reader (BioTek Instruments, Winooski, VT). Data were analyzed from three independent experiments performed in triplicate, and non-linear regression and sigmoidal dose-response curves (GraphPad Prism, La Jolla, CA) were used to calculate IC₅₀ and R2 values.

All reagents for compound synthesis were used as supplied, and all solvents were ACS grade or better. Anhydrous solvents were stored over activated 4Å molecular sieves (Sigma Aldrich, 208604–1KG). The starting material 6-OH-BTA-1 (PiB) was obtained from ABX Biochemicals (Radeberg, Germany). The reagents propargyl-PEG₃-bromide (BP-22738) and 5/6-carboxyrhodamine 110-PEG₃-azide (BP-22478) were obtained from BroadPharm Inc. (San Diego, California).
Silica gel (pore size 60 Å, 200–400 mesh particle size) for column chromatography was obtained from Sigma Aldrich (Sigma Aldrich, 288549–500G). Analytical thin-layer chromatography was performed using glass-backed silica gel 60 F₂₅₄ TLC plates (Millipore, 1057150001) and aluminum-backed C18-W (RP-18W) silica F₂₅₄ TLC plates (Sorbtech, 2733167). Preparative thin-layer chromatography (pTLC) was performed using glass-backed C18-W (RP-18W) silica F₂₅₄ pTLC plates (Sorbtech, 2717124). Before use, pTLC plates were pre-washed with an overnight immersion in methanol followed by developing twice in methanol. The pTLC plates were then air-dried and activated at 120 ™C for 2 hours, after which they were stored in a desiccator until use. All 96-well plate-based fluorescence intensity measurements were recorded using a Biotek Synergy 4 plate reader (BioTek Instruments) using the xenon flash bulb.