QuantiChrom™ Acetoacetate Assay Kit

Protocol SDS

Application

• Direct assay of acetoacetate in serum, plasma, urine and other biological samples.

Key Features

• Sensitive and accurate. Linear detection range of 0.012 to 1 mM (1.2 – 100 nmole/well) for acetoacetate in 96-well plate assay.
• Convenient. No sample pretreatment needed. The procedure involves adding a single working reagent and reading the optical density at room temperature in kinetic mode.
• High-throughput. Can be automated as a high-throughput 96-well plate assay for thousands of samples per day.

Method

• OD550nm

Samples

• Serum, plasma, urine, and other biological samples

Species

• All

Procedures

• 30 min

Size

• 100 tests in 96-well plate

Detection Limit

• 12 μM

Shelf Life

• 6 months

More Details

ACETOACETATE is one of three ketone bodies produced in the liver mainly from oxidation of fatty acids and are normally present at low concentrations in urine and blood. Increased ketone concentrations in the blood may lead to metabolic acidosis, which has been associated with diabetes, childhood hypoglycemia, growth hormone deficiency, alcohol or salicylate intoxication and inborn errors of metabolism. Acetoacetate is an unstable ketone body that spontaneously decarboxylates to release carbon dioxide and acetone. Simple, direct and automation-ready procedures for measuring acetoacetate (AAT) are very desirable. BioAssay Systems’ QuantiChrom™ Acetoacetate assay is based on a reaction with Sodium Nitroprusside forming a product with absorbance at 550 nm.

How stable is Acetoacetate?

Acetoacetate is an unstable ketone body that spontaneously breaks down to acetone and carbon dioxide, or is converted to 3-hydroxybutyrate, which is more stable. Because of this inherent instability, samples should be assayed fresh, or stored at -80°C and assayed shortly thereafter.

Do samples need to be deproteinated?

This kit can measure acetoacetate in the presence of proteins, but there is the potential for enzymatic conversion of acetoacetate to 3-hydroxybutyrate. To deproteinate samples, pass the sample through a 10 kDa NMWCO spin filter. For low levels of acetoacetate or in cases where you suspect matrix interference, it may be necessary to employ an acetoacetate spike, or internal standard, in the sample.

What is the normal concentration of acetoacetate in the blood?

The normal levels of acetoacetate in non-diabetic patients should be below 66 μM, but the levels of acetoacetate in a patient experiencing diabetic ketoacidosis can be considerably higher. Please keep in mind that this kit is for research use only, and not to be used for diagnostic purposes.

For more detailed product information and questions, please feel free to email or call us at 1-510-782-9988 x 2. For more general information regarding our assays, please refer to Technical Support.

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If you or your labs do not have the equipment or scientists necessary to run this assay, BioAssay Systems can perform the service for you.

– Fast turnaround
– Quality data
– Low cost

Please email or call 1-510-782-9988 x 2 to discuss your projects.