About 15,000 primary mouse hepatocytes (Lonza) per well were seeded in BioCoat Collagen I 96-well flat-bottom plates (catalog no.: A11428-03; Thermo Fisher Scientific) in 100 μl Hepatocyte Plating Medium (catalog no.: MP250; Lonza). Supernatant was removed 4–6 h after seeding, and ASOs were added at indicated concentrations (5,000, 1,000, 200, 40, 8, and 1.6 nM) diluted in 100 μl Maintenance Medium (catalog no.: MM250; Lonza). Cells were cultured for 3 days at 37°C. Afterward, the cells were lysed, and mRNA levels were measured according to the manufacturer's instructions, using the QuantiGene Singleplex Gene Expression Assay (catalog no.: QS0011; Thermo Fisher Scientific) and the following probesets: murine ANGPTL4 (catalog no.: SB-16744-02; Thermo Fisher Scientific) and HPRT1 (catalog no.: SB-15463; Thermo Fisher Scientific). Values were normalized to the housekeeping gene HPRT1. IC50 values were calculated using Prism 6 (GraphPad Software, Inc). Data are represented as the mean of triplicate wells ± SD relative to untreated cells (set as 1).
Hepatocyte plating medium
Manufactured by Lonza
Sourced in United States
Hepatocyte plating medium is a cell culture medium designed to support the growth and maintenance of primary human hepatocytes. It provides the necessary nutrients and growth factors to facilitate the attachment and initial culture of these specialized liver cells.
Automatically generated - may contain errors
Lab products found in correlation
Hprt1, by Thermo Fisher Scientific (1 mentions)
Prism 6, by GraphPad (1 mentions)
Hbm basal medium, by Lonza (1 mentions)
Hek293, by American Type Culture Collection (1 mentions)
Rat c6 glioma cells, by American Type Culture Collection (1 mentions)
Accutase, by STEMCELL (1 mentions)
Maintenance medium, by Lonza (1 mentions)
Matrigel, by Lonza (1 mentions)
Collagen coated 96 well plates, by Corning (1 mentions)
4 protocols using hepatocyte plating medium
1
Quantifying Mouse Hepatocyte ANGPTL4 mRNA
2
Culturing Diverse Cell Lines for Research
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Human pancreatic islets were obtained from the Integrated Islet Distribution Program (IIDP) funded by the National Institute of Diabetes and Digestive and Kidney Disease (NIDDK). Islets were cultured in low-glucose-containing CMRL1066 medium with supplements as described in suspension culture dish.17 (link) HEK293 (ATCC), C6 rat glioma cells (ATCC), and PD220 primary human fibroblast cells (OHSU Fanconi Anemia Research Materials) were cultured in DMEM with high glucose supplemented with 10% fetal bovine serum (FBS). MIN6 mouse insulinoma (ATCC) cells were grown in DMEM with high glucose and 15% FBS and αTC1 mouse clone 6 alpha cells (ATCC) in DMEM with low glucose (1 g/L), 15% HEPES, and nonessential amino acid. Primary human hepatocytes were plated in hepatocyte plating medium (Lonza) at a density of 80,000 cells/cm2, and media were changed to culture media consisting of HBM basal medium and HCM supplement (Lonza) 18 h after the plating. SH-SY5Y human neuroblastoma (ATCC) cells were grown in culture media recommended by ATCC.
3
Primary Human Hepatocyte Isolation
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Metabolism qualified primary human hepatocyte was purchased from Lonza (Walkersville, MD). Cells were grown adherently in hepatocyte plating medium (Lonza). For transfection cells were dissociated with Accutase (StemCell Technologies) to single cells.
4
Cryopreserved Hepatocyte Culture Protocol
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Plateable and interaction-qualified cryopreserved human hepatocytes (Lonza, Walkersville, MD, USA) were thawed in a hepatocyte thawing medium (Lonza) and then seeded at 5 × 104 cells per well in collagen-coated 96-well plates (Corning, Corning, NY, USA) with hepatocyte plating medium (Lonza). After 1 h of incubation with 5% CO2 under saturated humidity at 37 °C, the medium was replaced with fresh plating medium. After 4 h of seeding, the media were replaced with maintenance medium (Lonza) containing Matrigel (0.3 mg/ml).
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