EXPERIMENT 6
ASSAY OF BLOOD SUGAR
Kit for assaying blood glucose (for hand and automatic analyzer)
(glucose oxidase—peroxidase)
INTRODUCTION
This kit is used to assaying the concentration of glucose in human serum.
Glucose is a major sugar in blood, which is the transporting form. The
concentrations of glucose will increase in some patients, who suffer from enrocrine
disease such as DM(diabetes mellitus),endocrine diseases (hyperthyroidism ，
hypercorticism ; And the concentration will be low in some long –term malnutrition
people or the men who can not eat. Starvation or violent exercise make the
concentration descent decline temporarily.
PRINCIPLE
Glucose + H2O + O2
Glucose oxidase
gluconic acid + H2O2
Peroxidase
H2O2 + 4-aminoantipyrine + phenol
The red compound of quinone + H2O
The absorption of the red compound of quinone will be measured at the
505nm with 721 spectrophotomery, and compare it with the absorption of the
standard glucose solution and calculate the concentration of the specimen.
MAJOR REAGENT
Enzyme reagent :
(10mL, Concentrated)
glucose oxidase
peroxidase
phosphoric acid buffer
4-aminoantipyrine
Phenol reagent(5mL,Concentrated) : phenol
Glucose standard solution :
5.55mmol/L
(Applicated enzyme reagent 100mL,phenol reagent 100mL,component is the
same as the concentrated)
STORAGE AND STABILITY
1. Type concentration reagent may store at 4℃～8℃ for one year.
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2. Type application reagent may store at 4℃～8℃ for one year.
3. The dispensed compound of enzyme-quinone may store 4℃～8℃ for one
month.
COLLECTION OF SPECIMEN
1. The specimen may be serum or plasm anti-coagulated by heparin or EDTA.
2. The serum or plasm should be separated from the blood cells in one hour after
collection. If adding glycongen glycolysis inhibitor (NaF,KF),the serum or
plasm will be store at 15℃ ～25℃ for 24 hours.
PROCEDURE
The concentrated enzyme reagent should be diluted from 10mL to 100mL with
distilled water and phenol reagent should be done from 5mL to 100mL with distilled
water before assaying. Then, mix the two reagents for using.
If it is the applicated reagent, you may mix the enzyme reagent and phenol
reagent by 1∶1 for using.
STEPS OF ASSAYING BY HANDS
REAGENT
Glucose standard solution
(5.55mmol/L)
Serum / plasm
Distilled water
Compound of enzyme-phenol
STANDARD TUBE
10 μL
—
—
1.5mL
SPECIMEN TUBE
CONTROL TUBE
—
—
10 μL
—
1.5mL
—
10 μL
1.5mL
Mix the three tubes sufficiently, and put them into water both at 37℃ for 15
min .After adjusting zero with control tube the absorptions of tubes are measured at
505nm by with 721 spectrophotometry .
If using automatic biochemistry analyzer, please write program on analyzer’s
request.
CACULATION
Aspecimen
BLOOD SUGAR(mmol/L)=
× 5.55
Astandard
If the concentration of blood sugar > 22 mmol/L, please dilute the serum or
plasm firstly and the result of assaying should time times of dilution.
REFERENCE VALUE
2
Serum/plasm : 3.9～6.1mmol/L
PERFORMANCE INDICATOR
1. ACCURACY: the mean rate of recover is at 95.0%～105.0% at adding
glucose 2.8mmol/L～16.7mmol/L.
2. PRECISION : coefficient of variation(C.V.) within block <2%,among blocks
<3%.
3. ASSAYING RANGE : linear relation at 0～22mmol/L
(Translated by Yongjun Liu)
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