Automated kinetic method for D-3-hydroxybutyrate in plasma or serum

Area of Science:

• Clinical Chemistry
• Biochemical Assays

Background:

• Ketosis diagnosis relies on accurate metabolite measurement.
• Existing methods for 3-hydroxybutyrate analysis may have limitations.

Purpose of the Study:

• To optimize and validate a kinetic assay for 3-hydroxybutyrate in biological samples.
• To establish a reliable method for identifying ketosis.

Main Methods:

• Kinetic initial-rate technique optimized for pH, enzyme, and cofactor concentrations.
• Assay validated on Gilford 103 and Hitachi 705 analyzers.
• Interference from lactate dehydrogenase/lactate eliminated using oxalate.

Main Results:

• Optimized assay parameters: pH 8.5, hydroxybutyrate dehydrogenase 62.5 U/L, NAD+ 20 mmol/L.
• Assay demonstrated good agreement with a segmented-flow method.
• Within-assay precision ranged from 7.8% to 0.6%; analytical recovery was 91-99%.

Conclusions:

• The optimized kinetic assay provides accurate and precise measurement of 3-hydroxybutyrate.
• This method facilitates the identification of clinical and subclinical ketosis in humans and animals.
• Reagents are stable, enhancing practical application.