Cary eclipse varia spectrofluorometer

CFBE41o − cells were plated with density of 25 × 10³ cells/well in a 96 well plate with clear bottom (Coaster®, Corning, Germany) for 24 h [23 (link)–25 (link)]. Cells were then treated with different concentrations of lumacaftor alone (0.1 μM, 3 μM, 8 μM, 10 μM), ivacaftor alone (0.1 μM, 3 μM, 8 μM, 10 μM), a combination of the both (0.1 μM, 3 μM, 8 μM, 10 μM), and drugs–PEG–NLC (3 μM). After 48 h, all cells were loaded with 10 mM N-(Ethoxycarbonylmethyl)-6-Methoxyquinolinium Bromide (MQAE) for 2 h, then cells were washed with a chloride buffer containing of 140 mM NaCl, 5 mM HEPES, 5 mM KCl, 1 mM MgCl₂, 5 mM glucose, and 1 mM MgCl₂ pH 7.4, incubated with this buffer for 1 h at 37 °C with 5% CO₂ followed by exposure to a chloride-free buffer of similar composition (NO₃ as the substituting anion). After that, cells were exposed for 10 min to chloride-free buffer containing 100 mM IBMX and 5 mM forskolin (Sigma, Louis, MO). Fluorescence was recorded with a Cary Eclipse Varia spectrofluorometer (Varian, Walnut Creek, CA), using 360 nm (bandwidth 10 nm) as excitation wavelength and 450 nm (bandwidth 10 nm) as emission wavelength. All measurements were done in triplicate using the untreated CFBE41o- cells as a control.