Reader infinite f201

LDH levels in vascular and hepatic medium supernatants were measured by using the Cytotoxicity Detection Kit^PLUS (LDH) (Roche, Basel, Switzerland). Therefore, samples were collected from drug-treated liver models every 24 h for up to 72 h. Supernatants were directly diluted 1:2 in LDH storage buffer containing 200 mM Tris (Carl Roth)-HCL (VWR International, Darmstadt, Germany) at pH 7.5, 10% glycerol (Sigma-Aldrich), 1% bovine serum albumin (Sigma-Aldrich) and AQUA AD iniectabilia to conserve LDH activity during freezing. Samples were stored at − 20 °C until performing the assay. Samples were thawed and the assay was performed according to the manufacturer’s instruction. LDH activity was analyzed spectrophotometrically at 490 nm and concentrations were calculated from a standard curve obtained from dilutions of the LDH standard (Tecan Reader infinite F201, Tecan).
The release of ALT into hepatic and vascular culture supernatants was analyzed using the Human ALT ELISA Kit (Abcam). The assay was performed according to the manufacturer’s protocol in a 384-well format with sample and reagent volumes accordingly adopted. ALT levels were determined spectrophotometrically at 450 nm and concentrations were calculated from an ALT standard curve (Tecan Reader infinite F201, Tecan).

Cellular viability of treated liver models was measured after 7 days of drug perfusion using the CellTiter-Glo Luminescent Cell Viability Assay (Promega, Madison, WI, USA). Biochip membranes were excised and transferred to a 48-well microplate containing cell-specific phenol red-free William’s Medium E medium. CellTiter-Glo Reagent was added at a ratio of 1:1. The microplate was shaken for 2 min on an orbital plate shaker and was afterwards incubated for 10 min at RT. The solution was transferred to a white 96-well microplate. The luminescence signal was measured in a luminescence plate reader (Tecan Reader infinite F201, Tecan, Männedorf, Switzerland).