Total ketone bodies acetoacetate AcAc 3-hydroxybutyrate 3-HB enzymatic assay R1 3-1000µmol/L Standard 0.2-200µmol/L High-Sens

Total Ketone Bodies Assay R1 manufactured by FUJIFILM Wako Pure Chemical Corporation

Glucose is the preferred energy source for cells when this is not available the body will switch to fat metabolism to produce energy. Ketones are a product of fat metabolism and levels of ketones will build up in the blood, firstly producing ketosis and then progressing to ketoacidosis. Ketoacidosis is a form of metabolic acidosis most often seen is individuals with uncontrolled type I diabetes, although it can occur in type II diabetes, during pregnancy, illness and severe infections. There are three ketone bodies - acetoacetate, acetone and beta-hydroxybutyrate (a.k.a. 3-HB). Beta-hydroxybutyrate (3-Hydroxybutyrate, 3-HB) is the main ketone present in severe diabetic ketoacidosis and measurement of 3-HB and Total Ketone bodies can be useful in the diagnosis and monitoring of diabetes. The AcAc/3-HB ratio in arterial blood reflects liver reserve capacity so may also be useful in the field of liver transplantation.

The Wako Total Ketone Bodies assay measures both acetoacetate (AcAc) and 3-Hydroxybutyrate (3-HB, also known as beta-hydroxybutyrate). When used in conjunction with the 3-HB assay, the concentration of AcAc can be calculated by subtracting the 3-HB value from the Total value. The Total Ketone Bodies assay is performed using the Total Ketone Bodies Reagent (415-77301) and the 3-HB Reagent 2 (413-73601). When the sample is mixed with these reagents a cyclic reaction is triggered whereby AcAc is reduced to give 3-HB (oxidation of NADH to NAD) and 3-HB is oxidised to give AcAc (reduction of Thio-NAD to Thio-NADH). Both AcAc and 3-HB are assayed by measuring the rate of Thio-NADH production at 405nm. Wako's Total Ketone Bodies assay can be used with serum or plasma (heparin, citrate oxalate, EDTA or sodium fluoride) samples and has a standard measuring range of 3-1000µmol/L (a high-sensitivity method using increased sample gives a range of 0.2-200µmol/L).

The assay utilises two reagents: Total Ketone Bodies Reagent 1 (415-73301) is prepared by reconstituting the lyophilised Thio-NAD with phosphate buffer, this solution is stable for up to 3 weeks when stored at 2-10C. 3-HB Reagent 2 (417-73501) is prepared by combining lyophilised 3-HBDH and NADH with Good's Buffer; the reconstituted reagent is stable for up to 3 weeks when stored at 2-10C. The Total Ketone Bodies assay should be calibrator with the Ketone Body Calibrator (412-73791). 3-HB can be determined by using 3-HB Reagent 1 (417-73501) with 3-HB Reagent 2 (417-73501) and AcAc calculated by subtracting 3-HB from Total Ketones.