Sudan Red III in situ staining of cultured primary rat hepatocytes
Scope of the method
- Human health
- Basic Research
- Translational - Applied Research
- In vitro - Ex vivo
- Animal derived cells / tissues / organs
- Formaldehyde fixation
- Sudan Red III staining
- Hematoxylin nuclear counterstain
- Primary rat hepatocytes
- Intracellular lipids
- in vitro
- Drug-induced cytotoxicity
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 ability of a lysochrome, i.e. Sudan Red III diazo-dye to stain intracellular lipids. Additionally, subsequent application of hemalum, which is a complex formed by aluminium ions and oxidized haematoxylin, colours nuclei of the cells and thus enables their localisation. Red-coloured lipid droplets and blue nuclei are readily visible upon examination of the cells under a light microscope.
- Inverse-phase light microscope (Nikon Optiphot) ;
- Oven (Thermo electron corporation, Heraeus, 60°C).
Pros, cons & Future potential
The standard operating procedure for Sudan Red III in situ staining of cultured primary rat hepatocytes is easily applicable and allows a simultaneous screening of multiple compounds and/or multiple concentrations of the same compounds (to examine chemically induced steatosis).
Sudan Red III stain has a high affinity to a broad range of lipids and consequently does not discriminate between e.g. neutral lipids and phospholipids. Therefore, it is of utmost importance to perform more than one assay or use a more specific assay.
References, associated documents and other information
Contact personRaf Van Campenhout
OrganisationsVrije Universiteit Brussel
Pharmaceutical and Pharmacological Sciences (FARM)
In Vitro Toxicology and Dermato-cosmetology