- Provides a deeper understanding of ketone bodies than ketone body assays that only measure beta hydroxybutyrate.
- FuncS
Supplier Data
Functional Studies - Ketone Body Assay Kit (AB318308)
Recovery of β-Hydroxybutyrate and Acetoacetate spiked into pooled human plasma
β-Hydroxybutyrate and Acetoacetate in pooled human plasma (20 µL, 4 µL). Samples were spiked with β-Hydroxybutyrate (5 nmol) and Acetoacetate (20 nmol), respectively. The average spike recoveries were 83% for β-Hydroxybutyrate and 92% for Acetoacetate.
- FuncS
Supplier Data
Functional Studies - Ketone Body Assay Kit (AB318308)
Acetoacetate Standard Curve. Acetoacetate concentration is directly proportional to the decrease in absorbance measured at 450 nm versus the reagent blank, hence the standard curve is graphed as the net difference in OD450 versus the reagent blank (|∆OD450|).
- FuncS
Supplier Data
Functional Studies - Ketone Body Assay Kit (AB318308)
β-Hydroxybutyrate Standard Curve.
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제품 세부 정보
Ketone Body Assay Kit ab318308 provides a combined kit to quantify total Ketone Body concentration by individually measuring both beta hydroxybutyrate and acetoacetate.
The term 'Ketone bodies' refers to three chemicals, acetoacetate, beta hydroxybutyrate and acetone. Ketone bodies are produced by the liver and used peripherally as an energy source when glucose is not readily available. The two main ketone bodies are acetoacetate (AcAc) and 3-β-hydroxybutyrate (3HB), while acetone is the third, and least abundant, ketone body.
Ketone body assay principle
The ketone body assay is based upon the beta hydroxybutyrate dehydrogenase-catalyzed oxidation/reduction of β-Hydroxybutyrate and Acetoacetate that selectively occurs at two different pH conditions, 9.5 and 6.5, respectively.
For the measurement of beta hydroxybutyrate, a pH 9.5 buffer is used, driving the enzyme to act on beta hydroxybutyrate, and reduce NAD+ to NADH. This subsequently reduces a colorimetric probe to produce a stable chromophore with an absorbance peak of 450 nm.
For the measurement of acetoacetate, a pH 6.5 buffer is used, driving the enzyme to act on acetoacetate, and oxidise NADH to NAD+. This results in a decrease in absorbance at 450 nm.
Getting the best performance from ab318308
Serum and plasma samples should be non-hemolyzed and processed immediately after collection. Acetoacetate is unstable and will rapidly degrade to acetone and CO2 at room temperature. We recommend analyzing serum/plasma samples promptly after collection.
If immediate assay is not possible, store samples at -80 °C (for up to 4 weeks). Prior to experiment, centrifuge serum/plasma at 14,000 x g and 4 °C for 10 min and transfer the supernatant to a new microfuge tube.
Keep all samples on ice during the assay.
Related and recommended products
beta Hydroxybutyrate (beta HB) Assay Kit ab83390 uses an identical method to this assay kit for the measurement of the beta hydroxybutyrate element of ketone bodies.
Acetoacetate Assay Kit ab180875 uses the non-enzymatic nitroferricyanide method to measure the acetoacetate element of ketone bodies.
The LOD for Beta-Hydroxybutyrate is 0.86 nmoles/well.
The LOD for Acetoacetate is 5.87 nmoles/well.
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This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
Acetoacetic acid functions as an essential intermediate in energy metabolism. Cells utilize it as an alternative energy source especially when glucose availability is limited. It does not act within a complex but plays an important role in the conversion of fatty acids to ketone bodies in the liver. The brain heart and skeletal muscles efficiently use ketone bodies during states of starvation or carbohydrate restriction.
Pathways
Acetoacetic acid participates mainly in the ketogenesis and ketolysis pathways. These processes involve the conversion of fatty acids to ketone bodies in the liver and the subsequent breakdown to produce energy in peripheral tissues. Acetoacetic acid is closely related to acetyl-CoA during these reactions reflecting its pivotal involvement in energy homeostasis when glucose supplies are scarce.
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