SERUM SGOT
SUMMERY:-
Serum Glutamate Oxaloacetate Transaminase (SGOT), also called as Aspartate aminotransferase (AST), belongs to the transferase class of enzymes. This enzyme shows high levels of activity in the heart, liver, skeletal muscles and kidneys. Since its level seems to be increasing enormously following Myocardial Infarction (MI), it can be used as supporting evidence in the diagnosis of MI (Especially 20-36 hrs after MI). Elevated levels are also seen in Viral / Toxic Hepatitis, Hepatic and Cardiac Necrosis, Muscular Dystrophy and Pulmonary Embolism.
AIM:-
Determination of Serum Glutamate Pyruvate Transaminase (SGPT or ALT) by Reifman & Frankel’s method
PRINCIPLE:-
Alanine aminotransferase (GPT) catalyzes the transfer of the amino group from alanine to oxoglutarate with the formation of glutamate and pyruvate.
L-Aspartate + 2-Oxogluatarate GPT Oxaloacetate +L-Glutamate
Oxaloacetate + 2,4 DNPH Brownish red colored complex
SGOT (AST) catalyses the transfer of amino group from Aspartic acid to 2- Oxoglutarate to form Oxaloacetate and L-Glutamate. The Oxaloacetate thus formed reacts with 2,4 Dinitrophenyl Hydrazine (2,4 DNPH) to form a corresponding Hydrazone, a brownish red colored complex in an alkaline medium.
The color intensity is directly proportional to the SGOT concentration in the serum and is measured photometerically at 505 nm (490- 546).
REQUIREMENTS:-
· Four test-tube,
· Colorimeter,
· SGOT Reagent Kit,
· Incubator,
· Cuvette,
· Pipette etc.
· Fresh Serum
PROCEDURE:-
1. Pipette into test tubes labeled as Blank, Calibrator, Control, Test and proceed as per given below
|
|
Blank |
Calibrator |
Control |
Test |
|
Substrate Reagent |
0.25 ml |
0.25 ml |
0.25 ml |
0.25 ml |
|
Deionized Water |
50µl |
|
|
|
|
Serum Sample |
|
|
|
50µl |
|
Calibrator (Conc. 170 U/L) |
|
50µl |
|
|
1. Mix and incubate at 37ºC for 60 minutes.
|
Color Reagent |
0.25 ml |
0.25 ml |
0.25 ml |
0.25 ml |
|
Serum Sample (Same Serum Sample Which is |
|
|
0.05 ml |
|
1. Mix and incubate at 37ºC for 20 minutes.
|
Alkaline Reagent (Prediluted) |
1.5 ml |
1.5 ml |
1.5 ml |
1.5 ml |
1. Read absorbance of all the tubes against distilled water at 505 nm
(490-546).
Calculation:-:-
SGOT (AST) activity in U/L = Abs of Test – Abs of Control x Conc of Calibrator (Conc :160 U/L)
Abs of Calibrator– Abs of Blank Then,
If Absorption of, Test = 0.35
Control = 0.30
Calibrator = 0.36
Blank = 0.05
Then,
SGOT (AST) activity in U/L = Abs of Test – Abs of Control xConc of Calibrator (Conc :160 U/L)
Abs of Calibrator– Abs of Blank Then,
SGOT (AST) activity in U/L = 0.35 –0.30 x 160 U
0.36 – 0.5
SGOT(AST) activity in U/L = 25.8
Result (U/L) = 25.8
NORMAL VALUE:- = 0-35 U/L
CLINICAL SIGNFINANCE
The group of
enzymes called transaminase exist in tissues of many organs. Necrotic activity
in these organs causes a release of measured. Since heart tissue is rich in AST
increased serum levels appear in patients after myocardial infraction, as well
as in patients with muscle disease. Muscular dystrophy and dermatomyositis .
The liver is especially rich in ALT, being this enzyme measurement used
primarily as a test for infectious and toxic hepatitis, although high levels of
both ALT and AST may also be found in cases of liver cell damage and acute
pancreatitis, suggesting that the obstruction of billary tree by the adematous
pancreas and the presence of associate hepatic disease may contribute to
elevated AST levels these patients. Slight or moderate elevations of AST and
ALT activities may be observed after intake of alcohol and after administration
of various drugs, such as salicylates, opiates, and ampicilin