The lipolysis assay is used to quantify glycerol that is released, along with free fatty acids, from cells during lipolysis. The glycerol and free fatty acids are formed in lipolysis when triglycerides are hydrolyzed.
Lipolysis Assay Kit ab185433 includes:
- the reagents to perform a glycerol assay, with a readout on a colorimetric plate reader
- washing and incubation buffers to use with cultured cells during a lipolysis treatment study
- isoproterenol to use as a positive control to stimulate lipolysis
How the lipolysis assay works
In the lipolysis assay protocol, cultured cells are washed with the supplied buffer, and then incubated either with the incubation buffer and isoproterenol, or in the conditions of your choice. Cell culture media is then collected for the glycerol assay.
In the glycerol assay, glycerol kinase phosphorylates glycerol; and then glycerol phosphate oxidase oxidizes glycerol-1-phosphate to produce dihydroxyacetone phosphate and hydrogen peroxide. The hydrogen peroxide reacts with a probe via a peroxidase to generate color (absorbance λ= 570 nm). The increase in absorbance is proportional to the amount of glycerol in the sample.
Lipolysis assay protocol summary:
- Wash cultured cells
- Incubate cells to stimulate lipolysis
- Transfer cell culture media to 96-well plate
- Add reaction mix
- Incubate for 30 min at room temperature
- Analyze with microplate reader
Related and recommended products
For fluorometric detection, we recommend Lipolysis Assay Kit (Fluorometric) Lipolysis Assay Kit (Fluorometric)ab185434
The glycerol assay used in this kit, is also sold separately as Free Glycerol Assay Kit Free Glycerol Assay Kitab65337.
Other notes
This product is manufactured by BioVision, an Abcam company and was previously called K577 Lipolysis (3T3-L1) Colorimetric Assay Kit. K577-100 is the same size as the 100 test size of ab185433.
Lipolysis is the hydrolysis of triglycerides within the cell into glycerol and free fatty acids. The glycerol and free fatty acids are then released into the bloodstream or culture media. Lipolysis occurs in essentially all cells, but is most abundant in white and brown adipose tissue. Deficiencies in lipolysis lead to increased intracellular lipid accumulation, resulting in abnormal cellular physiology, hyperlipidemia, and insulin resistance. Lipolysis can be induced by catecholamine and certain hormones. The kit includes synthetic catecholamine, Isoproterenol, which activates ß-adrenergic receptors. This leads to activation of adenylate cyclase, which catalyzes the conversion of ATP to cAMP. cAMP then serves as a second messenger to activate hormone-sensitive lipase, which hydrolyzes the triglycerides. This pathway can be inhibited by insulin.
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