Key features and details
- Assay type: Cell-based
- Detection method: Colorimetric
- Platform: Microplate reader
- Sample type: Adherent cells
Product nameHepatic Lipid Accumulation/ Steatosis Assay Kit
Sample typeAdherent cells
Species reactivityReacts with: Mammals, Other species
Hepatic Lipid Accumulation/ Steatosis Assay Kit (ab133131) provides a convenient tool for evaluating steatosis risk of drug candidates using Oil Red O to stain neutral lipids in hepatocytes. Lipid accumulation can be quantified using a plate reader after the dye is extracted from the lipid droplets. Chloroquine is included in the kit as a positive control for screening pharmaceutical candidates that induce steatosis in hepatocytes.
Steatosis, also known as fatty liver, is a pathological process characterized by abnormal accumulation of lipid within cells. There are two distinct patterns of steatosis: macrovesicular and microvesicular. The former is frequently seen in alcohol-induced liver injury, as a complication of metabolic syndrome such as obesity and type II diabetes, and is a marker of the hepatotoxic side effect of certain drugs. Microvesicular steatosis is more commonly related to mitochondrial dysfunction and defects in b-oxidation responsible for fatty liver seen in pregnancy and Reye’s syndrome. While simple steatosis may not be associated with significant impairment of liver function, extensive fat accumulation can lead to cirrhosis and even liver failure. Studies on alcohol-induced steatosis revealed a set of transcription factors which are thought to be involved in the process, including SREBP1, PPARa, and Erg-1. The mechanism of non-alcoholic steatosis formation is poorly understood and little information is available on the pathway(s) responsible for progressive hepatocellular damage following lipid accumulation.
Determining the hepatotoxicity of drug candidates is an essential component of the pharmaceutical discovery process. Steatosis is one of the parameters that is evaluated when determining the hepatoxicity of a drug candidate. In vitro liver models, such as the HepG2 cell line, are available for mechanism-based testing of the hepatotoxic effects of drug candidates. Chloroquine is a well known cationic amphiphilic drug that is known to induce steatosis which is often used as a positive control in drug screening for steatosis.
Storage instructionsPlease refer to protocols.
Components 1 kit Chloroquine Positive Control (25 mM) 1 x 50µl Fixative (10X) 1 x 10ml Lipid Droplets Assay Dye Extraction Solution 1 x 30ml Lipid Droplets Assay Oil Red O Solution 1 x 25ml Lipid Droplets Assay Wash Solution 6 x 30ml Steatosis Assay Hematoxylin 1 x 30ml
- Pathways and Processes
- Metabolic signaling pathways
- Lipid and lipoprotein metabolism
- Lipid metabolism
HepG2 cells were seeded at a density of 104 cells/well in a 96-well plate and grown in a 37°C incubator overnight. The next day, cells were treated with vehicle or different doses of chloroquine for three days. Left panel: HepG2 cells treated with vehicle. There are a few lipid droplets in the cells, appearing as red dots. Right panel: HepG2 cells treated with 25 µM chloroquine show significant accumulation of lipid droplets, evident by abundant appearance of red dots visualized by Oil Red O staining.
HepG2 cells were seeded at a density of 104 cells/well in a 96-well plate and grown in a 37°C incubator overnight. The next day, cells were treated with vehicle or different doses of chloroquine for three days. At the end of this incubation, cells were stained with Oil Red O according to the product protocol. Lipid accumulation was assessed by extracting the Oil Red O and measuring the absorbance at 490 nm.
ab133131 has been referenced in 3 publications.
- Holmgren G et al. Characterization of Human Induced Pluripotent Stem Cell-Derived Hepatocytes with Mature Features and Potential for Modeling Metabolic Diseases. Int J Mol Sci 21:N/A (2020). PubMed: 31940797
- Molina E et al. A Novel Y-Specific Long Non-Coding RNA Associated with Cellular Lipid Accumulation in HepG2 cells and Atherosclerosis-related Genes. Sci Rep 7:16710 (2017). Functional Studies ; Human . PubMed: 29196750
- Ren T et al. Dietary blueberry and bifidobacteria attenuate nonalcoholic fatty liver disease in rats by affecting SIRT1-mediated signaling pathway. Oxid Med Cell Longev 2014:469059 (2014). PubMed: 25544867