PROCEDURE : For liver function tests : SGPT and SGOT :
1. Preparation of standard graph for GOT / GPT assay :
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1 |
2 |
3 |
4 |
Blank |
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GOT / GPT substrate (ml) |
0.45 |
0.4 |
0.35 |
0.3 |
0.5 |
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Std. Sodium pyruvate (ml) |
0.05 |
0.1 |
0.15 |
0.2 |
- |
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Distilled water (ml) |
0.1 |
0.1 |
0.1 |
0.1 |
0.1 |
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DNPH reagent (ml) |
0.5 |
0.5 |
0.5 |
0.5 |
0.5 |
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Mix thoroughly, keep at room temperature for 20 minutes. |
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0.4N NaOH. (ml) |
5 |
5 |
5 |
5 |
5 |
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Karmen units (GOT) |
27 |
61 |
114 |
190 |
0 |
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Karmen units (GPT) |
28 |
57 |
97 |
150 |
0 |
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Mix and keep at room temperature for 10 minutes. Read absorbance at 530 nm. |
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A 530 GOT |
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A 530 GPT |
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Draw a curve of Karmen units on X axis versus absorbance on Y axis
2. Treatment of test sample :
• To 0.5 ml of GOT / GPT substrate prewarmed to 37°C, add 0.1 ml of serum and incubate at 37°C for 60 minutes for GOT / 30 minutes for GPT.
• Add 0.5 ml of DNPH. Mix thoroughly, keep at room temperature for 20 minutes.
• Add 5 ml of 0.4N NaOH. Mix and keep at room temperature for 10 minutes.
• Read absorbance at 530 nm.
Normal value :
The adult reference range for both AST and ALT is roughly 10-40 U/L i.e. 20.83 - 83.33 Karmen Units when measured at 37°C.
Although men have slightly higher values than women do, most laboratories use a single range for both genders.
Note :
• Karmen unit/ ml is equivalent to 0.48 U/L.
• The GOT / GPT substrate can be preserved in refrigerator by adding 1 ml of chloroform. Discard the substrate solution on appearance of turbidity.
