Glycerol Kinase
PREPARATION and SPECIFICATION
PREPARATION and SPECIFICATION
APPLICATIONS
This enzyme is used for diagnostic tests for the determination of triglycerides together with Glycerol-3-phosphate Oxidase

UNIT DEFINITION
One unit is defined as the amount of enzyme which consumes 1 μmol of glycerol per minute at 37 ℃ under the conditions specified in the assay procedure.
REAGENTS PREPARATION
Reagent I: 0.3 M glycerol.
Reagent II: 1 kU/mL POD.
Reagent III: 50 mM TOOS.
Reagent IV: 50 mM 4-AA.
Reagent V: 20 U/mL L-α-glycerophosphate oxidase.
Reagent VI: 200 mM HEPES, contains 20 mM MgCl2 and 40 mM ATP (pH 7.9).
Reaction mixture:
Reagent I 1.67 mL
Reagent II 400 μL
Reagent III 3 mL
Reagent IV 3 mL
Reagent V 40 mL
Pure water Set to 100 mL
Enzyme dilution buffer: 20 mM, pH 7.5 potassium phosphate buffer.
Sample: Dilute the sample to 0.2 – 0.4 U/mL by enzyme dilution buffer.
PROCEDURE
- Add 1mL reaction mixture to the 1 mL cuvette.
- Preincubate at 37 ℃ for 2 min.
- Add 0.02 mL enzyme solution to the cuvette and mix.
- Record the ΔAs at 555 nm in 1 minute in a spectrophotometer thermo-stated at 37 ℃.
At the same time, measure the blank rate ΔAb by using the same method as the test except that the enzyme diluent is added instead of the enzyme solution.
∆A = ∆As – ∆Ab

REFERENCES
- Sakasegawa, S., et al. (1998) Biosci. Biotechnol. Biochem., 62, 2388-2395.