Omega mars data analysis software package

Cytotoxic effects of compounds used for the cell layer integrity study were evaluated by using a resazurin-based in vitro toxicology assay (Sigma-Aldrich; Schnelldorf, Germany), according to the manufacturer’s instructions. Briefly, cells were seeded into 96-well plates (45,000 cells per well), grown to 90% confluence, and incubated with the test substances for 2 h at 37 °C. The cells were washed and incubated with 10% resazurin in growth medium for 2 h. Subsequently, the amount of the reduced form of resazurin (resorufin) was determined with a microplate reader in fluorescence mode (544 nm excitation, 590 nm emission; POLARstar Omega, BMG LABTECH, Ortenberg, Germany). Data analysis was done by OmegaMARS Data analysis software package (BMG LABTECH, Ortenberg, Germany). Untreated cells grown under the same conditions were used for normalization of cell viability. Each test substance was measured in triplicate.

Cell viability was evaluated using a resazurin-based in vitro toxicology assay according to the manufacturer’s protocol. Briefly, cells were seeded in 96-well plates at 5 × 10⁴ cells per well, grown to 80% confluence and incubated with the indicated test substances at 37 °C for 2 h. Subsequently, the cells were washed and incubated with 10% resazurin in cell culture medium at 37 °C for 2 h. The level of the reduced form of resazurin (resorufin) was then determined using a microplate reader in fluorescence mode (544 nm excitation, 590 nm emission; POLARstar Omega, BMG LABTECH, Ortenberg, Germany). Data were analyzed using the OmegaMARS Data analysis software package (BMG LABTECH, Ortenberg, Germany). Cell viability was normalized to untreated cells grown under the same conditions. Each test substance was measured at least in quadruplicate.

The cytotoxic effects of herbal extracts used in this study were evaluated by using a resazurin-based in vitro toxicology assay (Sigma-Aldrich, Schnelldorf, Germany), according to the manufacturer’s instructions. Briefly, HuH7 and undifferentiated 3T3-L1 cells were seeded into 96-well plates (45,000 cells/well), grown to 90% confluency, and incubated with the test substances (BBR: 3.70 µg/mL; herbal extracts: 10 mg/L) for 1, 3 and 5 days at 37 °C. Subsequently, the cells were washed with medium and incubated with a medium containing 10% resazurin for 2 h. The concentrations of the reduced form of resazurin (resorufin) were then determined using a microplate reader in fluorescence mode (544 nm excitation, 590 nm emission; POLARstar Omega, BMG LABTECH, Ortenberg, Germany). Data were analyzed using the Omega MARS Data analysis software package (BMG LABTECH, Ortenberg, Germany). Cell viability was normalized to untreated cells grown under the same conditions. Each test substance was measured in triplicate. The same experiment was performed with differentiated 3T3-L1 adipocytes, as well as during differentiation (treatment started at medium 2 addition).

The cytotoxic effects of EOs used in this study were evaluated by using a resazurin-based in vitro toxicology assay (Sigma-Aldrich, Schnelldorf, Germany), according to the manufacturer’s instructions. Briefly, cells (Hela: 40,000, Caco-2: 120,000, STF1: 40,000 cells/well) were seeded into 96-well plates, grown to 90% confluency, and incubated with EOs at different concentrations (0.0016–1% [v/v]) for 24 h at 37 °C. Subsequently, the cells were washed and incubated with medium containing 10% resazurin for 2 h. The concentrations of the reduced form of resazurin (resorufin) were then determined using a microplate reader in fluorescence mode (544 nm excitation and 590 nm emission; POLARstar Omega, BMG LABTECH, Ortenberg, Germany). Data were analyzed using the Omega MARS Data analysis software package (BMG LABTECH, Ortenberg, Germany). Cell viability was normalized to untreated cells grown under the same conditions. Each test substance was measured in triplicate.

Cytotoxic effects of compounds under study were evaluated by using a resazurin‐based in vitro toxicology assay (Sigma‐Aldrich; Schnelldorf, Germany), according to the instructions of the manufacturer. Briefly, cells were seeded into 96‐well plates (30 000 cells per well), grown to 90% confluence, and incubated with the test substances for 4 h at 37 °C. Subsequently, the cells were washed and incubated with a medium containing 10% resazurin for 2 h. Levels of the reduced form of resazurin (resorufin) were then determined using a microplate reader in fluorescence mode (544 nm excitation, 590 nm emission; POLARstar Omega, BMG LABTECH, Ortenberg, Germany). Data were analyzed using OmegaMARS Data analysis software package (BMG LABTECH, Ortenberg, Germany). Cell viability was normalized to untreated cells grown under the same conditions. Each test substance was measured in triplicate.