Nd 1000 uv vis thermo fisher nanodrop device

To confirm the results of the qPCR reaction and their translation into functional protein, the activity of the studied cytochromes was tested. We estimated the activity of the Cyp1a1/Cyp1b1, Cyp1a2, and Cyp2b1 enzymes using the fluorometric ethoxyresorufin-O-deethylase (EROD), 7-methoxyresorufin O-demethylation (MROD), and pentoxyresorufm O-dealkylase (PROD) substrates, respectively. The fluorescence EROD, MROD, and PROD assays were performed according to the method proposed by Kennedy et al. (1993 (link)). For the assays, the cells were seeded on 12-well plates and initially cultured for 24 h. Measurement of the EROD, MROD, and PROD activity was performed after 24 or 48 h of exposure to 1 μM βNF, Les-2194, Les-3640, Les-5935, and Les-6166. To perform the EROD, MROD, and PROD assays, lysed cells were transferred into multiwell plates, and the fluorescent product resorufin was quantified within the wells with a fluorescence plate reader (FilterMax F5) at an excitation wavelength of 530 nm and an emission wavelength of 590 nm. The protein concentration was determined spectrophotometrically in triplicate for each sample at 280 nm using the ND/1000 UV/Vis Thermo Fisher NanoDrop device.

We estimated the activity of the CYP1A1/CYP1B1 enzymes using the fluorometric ethoxyresorufin-O-deethylase (EROD) substrate. The fluorescence EROD assay was performed according to the method proposed by Kennedy et al. (1993 (link)). Briefly, the cells were seeded on 12-well plates and initially cultured for 24 h. The EROD activity was measured after the 24-h exposure to 1 µM βNF, 1 µM CAY10464, and 1 µM TDBP-TAZTO or in co-treatment with TDBP-TAZTO and the AhR agonist and antagonist. To perform the EROD assay, lysed cells were transferred into multiwell plates, and the fluorescent product resorufin was quantified within the wells with a fluorescence plate reader (FilterMax F5) at an excitation wavelength of 530 nm and an emission wavelength of 590 nm. The protein concentration was determined spectrophotometrically in triplicate for each sample at 280 nm using the ND/1000 UV/Vis Thermo Fisher NanoDrop device.