Journal: Current protocols in toxicology

Article Title: Micropatterned Co-Cultures of Human Hepatocytes and Stromal Cells for the Assessment of Drug Clearance and Drug-Drug Interactions

doi: 10.1002/cptx.23

Figure Lengend Snippet: Compounds that have been used for in vitro CYP450 profiling studies in micropatterned cocultures (MPCCs) All substrates can be reconstituted in dimethyl sulfoxide (DMSO) and incubated in MPCCs at the indicated concentrations for 1 hour. The supernatants are then collected and stored at -80°C prior to LC-MS/MS analysis or luminescence analysis (for the CYP2C9-Glo and CYP3A4-Glo substrates from Promega).

Article Snippet: Materials list-behavior=simple prefix-word= mark-type=none max-label-size=0 96-well assay plates 1.5 mL microcentrifuge tubes Multichannel micropipettes with tips Human hepatocyte dosing medium (see recipe in ‘Reagents and Solutions’ section) CYP450 luminescent kits (Promega) CYP450 substrates reconstituted in dimethyl sulfoxide (DMSO) Human hepatocyte maintenance medium (see recipe in ‘Reagents and Solutions’ section) Human albumin ELISA kit (Bethyl Laboratories) Urea nitrogen test kit (Stanbio Laboratory) Spectrophotometer (ideally compatible with 96-well plate for higher throughput) Luminometer (ideally compatible with 96-well plates for higher throughput) Data analysis software (e.g. Microsoft Excel and GraphPad Prism) Protocol steps list-behavior=enumerated prefix-word= mark-type=decimal max-label-size=0 Dilute the substrate from the Promega CYP450 kit (i.e. luciferin-IPA to measure CYP3A4 activity) in human hepatocyte dosing medium at a concentration recommended by the manufacturer. list-behavior=simple prefix-word= mark-type=none max-label-size=0 Alternatively, cultures can be dosed with a CYP450 probe substrate that requires liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for quantification of metabolite produced ( see ).

Techniques: In Vitro, Incubation, Concentration Assay

Journal: Military Medical Research

Article Title: Direct conversion of human fibroblasts into dopaminergic neuron-like cells using small molecules and protein factors

doi: 10.1186/s40779-020-00284-2

Figure Lengend Snippet: Phenotypic characterization of maturation neuronal markers in cells undergone the induction process. a-c Immunostaining assay for the mature neuron markers MAP2, neuronal nuclei (NEUN), and synapsin 1 (SYN) after 14 days of chemical induction. Scale bars = 25 μm

Article Snippet: After 6–8 days of chemical induction, the induction medium was replaced with the neuronal maturation medium with 5 μmol/L forskolin, 1 μmol/L kenpaullone, 0.2 mmol/L L-ascorbic acid (MedChem Express, #HY-B0166A), 100 ng/ml SHH, 100 ng/ml FGF-8b, 20 ng/ml bFGF, 10 ng/ml brain-derived neurotrophic factor (BDNF), 10 ng/ml glial cell-derived neurotrophic factor (GDNF).

Techniques: Immunostaining