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Block 161st Practical workDepartment of Clinical Pathology

Faculty of Medicine

Universitas Gadjah Mada Yogyakarta

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Glucose (GOD-PAP)

Principle:

Glucose concentration is determined after

enzymatic oxidation by glucose oxidase. Thecolorimeteric indicator is quinoneimine, which is

generated from 4-aminoantipyrine and phenol

by hydrogen peroxide under the catalytic action

of peroxidase (Trinders reaction).

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Reaction

Glucose + O2

GODGluconic acid + H2O2

2 H2O2 +

4Aminoantipyrine

+ Phenol

PODQuinoneimine +

4 H2O

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Serum, heparinized plasma or EDTA plasma,spinal fluid

Separate at the latest 1h after blood collection

from cellular contents.

Samples

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Components and Concentration in the test

Phosphate buffer pH 7,5 250 mmol/L

Phenol 5 mmol/L

4-Aminoantipyrine 0,5 mmol/L

Glucose oxidase(GOD) 10 kU/L

Peroxidase (POD) 1 kU/LStandard:100 mg/dL (5,55 mmol/L)

Reagents

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10 UL water

Mix well

10 uL

sample/std

BlankSample orstandard

1000 UL

reagent

1000 UL

reagent

incubate 20 minutes, at 20-25C or 10 min at 37C. Readabsorbance against the blank within 60 minutes

Mix well

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Calculation

With standard or calibrator

Glucose (mg/dL) = A Sample x Conc. Standard/Cal (mg/dL) A Std/Cal

Conversion factor

Glucose (mg/dL) x 0.05551 = Glucose (mmol/L)

Normal range:

Fasting 70 - 100 mg/dL (whole blood)

70 - 115 mg/dL (serum)35 - 50 mg/dL (spinal fluid)

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Glucose Challenge Test

Objective: To know body response after glucose loading

To screen DM among pregnant woman

Principle: Patient body force to respond by glucose loading

and the blood glucose are monitored.

Abnormal result can be obtained after Indicating patient:

Pregnancy 2nd trimester

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Technical Requirement

Samples:

Serum or plasma (recommended)

No finger prick blood samples

Reagents:

Glucose 50g

Reagent kit for glucose assay (GOD-PAP method)

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Assay Procedure

Patient is not fasting before test

Patient is given 50 g glucose loading, dissolved

in 200 ml water. The glucose should be drank

in 5 minutes

The patients venous blood sample is taken

one hour after loading

The glucose concentration is measured from

the sample

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Interpretation

Positive: venous glucose concentration >140

mg/dL.

If the result is positive, the patient should be

tested further by oral glucose tolerance test

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Oral Glucose Tolerance Test

Objective: To know body response after glucose loading and

to screen DM among suspected person

Principle: Patient body force to respond by glucose loading

Blood glucose are monitored every hour for 3

hours. Blood glucose curve can show abnormal

pattern

Indicating patient:

Screening test for DM is in a doubt

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Technical Requirement

Samples:

Serum or plasma (recommended)

No finger prick blood samples

Reagents:

Glucose 75g (PERKENI), or 100g (ADA)

Reagent kit for glucose assay (GOD-PAP method)

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Assay Procedure

Patient should fast for 8-12 hours before test

In fasting condition, blood sample is taken

Patient is given 75g or 100g glucose loading,

dissolved in appropriate amount of water

The patients venous blood sample is taken

one hour, two hours and three hours after

loading

The glucose concentration is measured from

the sample

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Interpretation

Normal criteria (WHO, 2004):

Fasting blood glucose:

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OGTT Interpretation in Gestational DM

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