Erythrocyte transketolase, a new semi-automated method

Area of Science:

• Biochemistry
• Clinical Chemistry
• Enzymology

Background:

• Transketolase (TK) is a key enzyme in the pentose phosphate pathway.
• Erythrocyte transketolase activity is a sensitive indicator of thiamine status.
• Accurate measurement of TK activity is crucial for diagnosing thiamine deficiency.

Purpose of the Study:

• To develop and validate a novel enzymatic assay for determining transketolase activity in human erythrocytes.
• To assess the utility of the assay with and without thiamine pyrophosphate (TPP) stimulation.
• To establish reference ranges for erythrocyte transketolase activity.

Main Methods:

• Hemolysed erythrocytes were incubated with ribose-5-phosphate.
• Glucose-6-phosphate formation was quantified enzymatically after deproteinization.
• Measurements were performed using a centrifugal fast analyser.
• The method was validated against an established colorimetric assay.

Main Results:

• The proposed enzymatic method demonstrated high throughput and accuracy.
• A strong correlation (r = 0.89) was observed between the new method and a colorimetric method.
• Reference values for erythrocyte transketolase activity in blood were determined to be in the range of 32--101 U/l.

Conclusions:

• The developed centrifugal analyser-based enzymatic assay is a reliable and efficient method for measuring erythrocyte transketolase activity.
• This assay can be used to assess thiamine status by evaluating enzyme activity with and without TPP stimulation.
• The established reference ranges provide a benchmark for clinical interpretation.