3-hydroxybutyrate 3-HB beta-hydroxybutyrate enzymatic assay R2 serum plasma 3-1000µmol/L standard 0.2-200µmol/L High-Sens Wako

3-hydroxybutyrate (3-HB) R2 serum 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.

In the presence of NADH and Thio-NAD and the enzyme 3-hydroxybutyrate dehydrogenase (3-HBDH) will catalyse the reduction of AcAc in the sample to 3-HB (oxidation of NADH to NAD) and the oxidation of 3-HB to AcAc (reduction of Thio-NAD to Thio-NADH). Measuring the rate of Thio-NADH production at 405nm is then used to quantify the ketone bodies in the assay sample.

This reagent 3-HB R2 is supplied as 2 vials of lyophilised reagent and 2x9ml reconstitution buffer. The reagent is prepared by combining lyophilised 3-HBDH and NADH with Good's Buffer; once reconstituted the reagent is stable for up to 3 weeks when stored at 2-10C. Ketone body assays require two reagents, to measure Total Ketones use with Total Ketone Bodies R1 (415-73301) and for 3-HB this reagent is used with 3-HB R1(417-73501).

When measuring Total Ketone Bodies the Total Ketone Bodies Reagent 1 (415-73301) is used with 3-HB Reagent 2 (417-73501) and both AcAc and 3-HB within the sample are assayed. Standard range 3-1000µmol/L (a high-sensitivity method using increased sample gives a range of 0.2-200µmol/L). Reagents are stable for up to 3 weeks after reconstitution if stored at 2-10C.

In order to measure just 3-HB the 3-HB Reagent 1 (417-73501) is used together with 3-HB Reagent 2 (417-73501). The sample is initially treated with acetoacetate decarboxylase (AADC) which breaks down all the original AcAc in the sample so that once the cyclic reactions with 3-HBDH begin only 3-HB is assayed. Standard range 3-1000µmol/L (a high-sensitivity method using increased sample gives a range of 0.2-200µmol/L). Reagents are stable for up to 3 weeks after reconstitution if stored at 2-10C.

Both the 3-HB and Total Ketone Bodies assays should be calibrated with Ketone Body Calibrator (412-73791).