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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Flow-cytometric determination of neutral lipids

Using this method you can measure the (relative) lipid load in human skin-derived stem cells differentiated towards hepatic cells. This method could also be applied on other cell types (e.g. HepG2), since it is based on the following publication: "M. T. Donato et al., Chem. Biol. Interact. 181, 417

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

Contact: Joost Boeckmans
Organisation: Vrije Universiteit Brussel (VUB)
Status: Internally validated
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Adult skin stem cell-derived in vitro model of hepatic steatosis

Human skin-derived adult stem cells differentiated towards hepatic cells (hSKP-HPC) are used in this method (R. M. Rodrigues et al., Stem Cells Dev. 23, 44–55 (2014)). These cells are exposed to a cocktail of insulin and glucose at certain concentrations. After 24h of exposure, these cells exhibit a

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

Contact: Joost Boeckmans
Organisation: Vrije Universiteit Brussel (VUB)
Status: Still in development
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In vitro human stem cell-based model of non-alcoholic steatohepatitis

Non-alcoholic fatty liver disease (NAFLD) ranges from simple steatosis to severe, life-threatening non-alcoholic steatohepatitis (NASH). Steatosis is mostly asymptomatic and does not cause health complications. However, in 5-10% of the cases it proceeds to NASH in which hepatic inflammation occurs

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

Contact: Joost Boeckmans
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 reactive oxygen species in cultured primary rat hepatocytes

Basically, the standard operating procedure outlined in this document consists of the following steps, namely, (i) preparation of the cells for the DCFH-DA assay, (ii) DCFH-DA test procedure, (iii) processing of the results. Practical details are provided for each of these steps and are followed by

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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