Acetoacetate Assay Kit (Colorimetric) (ab180875)
Key features and details
- Assay type: Quantitative
- Detection method: Colorimetric
- Platform: Microplate reader
- Assay time: 25 min
- Sample type: Other biological fluids, Plasma, Serum, Urine, Whole Blood
- Sensitivity: 25 µM
Overview
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Product name
Acetoacetate Assay Kit (Colorimetric) -
Detection method
Colorimetric -
Sample type
Urine, Serum, Plasma, Other biological fluids, Whole Blood -
Assay type
Quantitative -
Sensitivity
Assay time
0h 25mSpecies reactivity
Reacts with: Mammals, Other speciesProduct overview
Acetoacetate Assay Kit (Colorimetric) ab180875 adapts the traditional qualitative assay using dipsticks, and sodium nitroferricyanide as a chromophore, with a modification that provides a sensitive method to quantitate endogenous levels of AcAc in human blood and urine.
In this non-enzymatic assay, AcAc reacts with a substrate to generate a colored product that can be measured at 550 nm. The reaction is specific for AcAc and does not detect 3-β-hydroxybutyrate. The assay kit can detect samples containing acetoacetate as low as 25 µM.
Acetoacetate assay protocol summary:
- add samples and standards to wells
- add reaction mix
- analyze with a microplate reader in kinetic mode for 10-15 minNotes
Acetoacetate (AcAc), a β-ketoacid, is one of the three ketone bodies and is formed via condensation of two molecules of acetyl-CoA in liver mitochondria. AcAc can be enzymatically reduced to 3-β-hydroxybutyrate (β-HB), or decarboxylated producing acetone ((CH3)2CO). Ketone bodies [βHB: 78%; AcAc: 20% & (CH3)2CO: 2%] are mainly used as an alternative energy source when glucose cannot be delivered to the system. Excessive concentration of ketone bodies (ketoacidosis) is observed in patients with Type I diabetes, severe starvation or alcoholism. Traditionally, AcAc levels have been qualitatively detected using dipsticks that use sodium nitroferricyanide as a chromophore.
Platform
Microplate readerProperties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components Identifier 100 tests Acetoacetate Assay Buffer 1 x 15ml Acetoacetate Standard (Lyophilized) Yellow 1 vial Acetoacetate Substrate 1 x 1ml
Properties
-
Storage instructions
Store at -20°C. Please refer to protocols. -
Components Identifier 100 tests Acetoacetate Assay Buffer 1 x 15ml Acetoacetate Standard (Lyophilized) Yellow 1 vial Acetoacetate Substrate 1 x 1ml
Images
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Functional assay - 180875 Acetoacetate Assay Kit (Colorimetric)
Acetoacetate measured in urine samples showing concentration (micromolar).
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Functional assay - 180875 Acetoacetate Assay Kit (Colorimetric)Colorimetric standard curve: mean of duplicates (+/-SD) with background readings subtracted.
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Measurement of Acetoacetate in urineSamples were deproteinized by spin filtering. 100 µL of undiluted urine was spiked with known amounts of AcAc (30 nmol).
Assays were performed following the kit protocol.
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Standard CurveStandard Curve for equimolar amounts of Acetoacetate, β-HB and Acetone (4°C, 100 min.). The other two ketone bodies β-hydroxybutyrate or Acetone are not detected in the assay due to much lower sensitivity (β-HB) and much lower concentration (Acetone).
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Time course experimentTime course of color development at 4°C and 25°C (100 nmol Acetoacetate).