Isolation of rat hepatocytes

This method describes the steps from a living rat to a single cell solution of primary hepatocytes. This requires surgery on the lab animal, a perfusion with buffer solution, a digestion with collagenase and a filtration step to obtain primary hepatocytes.

Last updated on: 28-02-2022 - 11:01

Contact: Paul Claes
Organisation: Vrije Universiteit Brussel (VUB)
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Establishment of sandwich cultures of primary human hepatocytes

This method describes a well-known optimised human in vitro model of drug-induced cholestasis. Cryopreserved primary human hepatocytes are cultured between two layers of extracellular matrix scaffold, which will delay dedifferentiation and allows to restore cell-extracellular matrix interactions.

Last updated on: 28-02-2022 - 11:00

Contact: Eva Gijbels
Organisation: Vrije Universiteit Brussel (VUB)
Status: Still in development
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Microfluidic perfusion culture for hepatic differentiation of human skin stem cells

The effect of fluidics that mimic the blood flow in the liver sinusoids, is evaluated during the hepatic differentiation of human skin-derived precursors (hSKP). In a standard bi-dimensional (2D) cell culture system, hSKP are differentiated to hSKP-HPC for 24 days in static conditions. In a

Last updated on: 28-02-2022 - 11:00

Contact: Alessandra Natale
Organisation: Vrije Universiteit Brussel (VUB)
Status: Still in development
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Manufacturing of fiber scaffolds and cell seeding

Polycaprolactone (PCL) fiber materials are fabricated using an electrospinning method. Molecular weight of PCL is 45 000 Da (Sigma-Aldrich). The electrospinning process is performed using 18wt% PCL solution dissolved in chloroform:ethanol at a ratio of 9:1. Two high-voltage sources are used to

Last updated on: 28-02-2022 - 11:00

Contact: Alessandra Natale
Organisation: Vrije Universiteit Brussel (VUB)
Status: History of use
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Spheroid cell culture of human skin-derived precursors

For the generation of the human skin-derived precursors (hSKP) spheroids, ultra-low attachment 96-well plates, cell-repellent (Greiner) are used. hSKP cultured on tissue culture plates are detached with trypple (5 minutes) and collected in a tube. After centrifugation, hSKP are counted at the Burker

Last updated on: 28-02-2022 - 10:59

Contact: Alessandra Natale
Organisation: Vrije Universiteit Brussel (VUB)
Status: Still in development
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Measurement of the extracellular release of adenosine triphosphate in cultured primary rat hepatocytes

ATP transports chemical energy within cells by serving as a substrate for kinases and as such fulfills a vital function in numerous cellular processes such as cell injury and subsequent cell death. ATP is therefore a crucial player in these events that are results of intracellular stress.

Last updated on: 28-02-2022 - 10:59

Contact: Axelle Cooreman
Organisation: Vrije Universiteit Brussel (VUB)
Status: History of use
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Cytotoxicity measurement in cultured primary rat hepatocytes

The MTT test is performed to determine the in vitro cytotoxicity of selected chemicals. The mitochondrial enzyme succinate deydrogenase is responsible for the biotransformation of toxic agents and MTT. The ability of cells to reduce MTT provides an indication of the mitochondrial integrity and

Last updated on: 28-02-2022 - 10:59

Contact: Axelle Cooreman
Organisation: Vrije Universiteit Brussel (VUB)
Status: History of use
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The detection of cholestasis-inducing agents in cultured primary rat hepatocytes

The standard operating procedure describes a method to assess the cholestasis-inducing potential of chemicals, in casu in cultures of primary rat hepatocytes. The procedure relies on the accumulation of the fluorescent bile salt export pump (Bsep) substrate cholyl-lysyl-fluorescein (CLF) in the

Last updated on: 28-02-2022 - 10:59

Contact: Raf Van Campenhout
Organisation: Vrije Universiteit Brussel (VUB)
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Sudan Red III in situ staining of cultured primary rat hepatocytes

The standard operating procedure for Sudan Red III in situ staining of cultured rat hepatocytes describes how to detect one of the aspects of drug-induced cytotoxicity i.e. the intracellular accumulation of lipids or in other words steatosis, in primary rat hepatocyte cultures. It is based on the

Last updated on: 28-02-2022 - 10:59

Contact: Raf Van Campenhout
Organisation: Vrije Universiteit Brussel (VUB)
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Measurement of urea synthesis in cultured stem cell-derived hepatocyte-like cells

The present standard procedure describes a protocol for measuring the urea concentration in supernatant of human stem cell-derived hepatocyte-like cells. This procedure relies on a chromogenic reagent that forms a colored complex specifically with urea. The latter can be measured and is directly

Last updated on: 28-02-2022 - 10:59

Contact: Cannot be disclosed
Organisation: Vrije Universiteit Brussel (VUB)
Status: History of use, Internally validated
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