Estimation of transaminase in serum as ALT , AST – liver function tests

ALT , AST test

    Liver Function Tests (LFT): are group of clinical biochemistry laboratory blood assays designed to give information about the state of patients’liver.
    Estimation of transaminase in serum as ALT , AST
    • Introduction
    ALT test :
    • ALT is an enzyme normally found inside liver cells 
    • The function of ALT is to convert alanine (A.As) into pyruvate
    • ALT is more spesific for the liver disease
    • In healthy individuals , ALT levels in the blood is low
    • When the liver is damaged , ALT is released into the blood usually before more signs of liver damage occur.this makes ALT a useful test for early detection of liver damage but it can’t show how much liver damage there is or how much fibrosis or scarring is present
    • AlT test may also performed to :
    1. Monitor the progression of liver diseases as hepatitis
    2. Assess whether treatment for liver disease should be started
    3. Evaluate how well treatment is working

    Estimation of transaminase in serum as ALT , AST

     AST test :
    • AST is an enzyme found in cells throughout the body but mostly in the heart , liver and to a lesser extent in the kidneys and muscles so, it is less specific for liver disease
    • The function of ALT is to catalyze the reversible transfer of an amino group between aspartate and glutamate
    • To catalyze the inter conversion of aspartate and 𝜶 – keto glutarate to oxaloacetate and glutamate
    •                                                                                         GPT
    • Alanine + 𝜶-  keto glutarate Pyruvate + Glutamate                                 
    •                                                                                               GoT            
    • Aspartate +𝜶-  keto glutarate Oxaloacetic acid + Glutamate
    • The pyruvate produced by transamination by GPT reacts with 2,4-dinitrophenyl hydrazine to give a brown colored complex (hydrazone) which is measured by the colorimeter at 505 nm
    • The oxaloacetate formed in the reaction with GOT decarboxylated spontaneously to pyruvate which converted to hydrazone also 
    • The transaminase activity is proportional to the amount of oxalate to pyruvate formed over a definite period of time and is measured by 2,4-dinitrophenyl hydrazine in alkaline solution

    Normal range
    • ALT (GPT) up to 45 units / ml
    • AST (GOT) up to 40 units / ml
    • Phosphate buffer :PH 7.5 (optimum PH of enzyme activity)
    • ASTsubstrate 
    • ALT substrate
    • Color reagent 
    • Standard pyruvate
    • NaoH 
    • Depend on colorimetric technique
    Dist H2o
    0.05 ml
    0.05 ml
    0.05 ml
    0.05 ml
    0.05 ml
    Alt or Ast substrate
    0.25 ml
    0.25 ml
    0.225 ml
    0.2 ml
    0.175 ml
    0.15 ml
    0.25 ml
    0.25 ml
    0.05 ml
    0.075 ml
    0.1 ml
    Color reagent
    0.25 ml
    0.25 ml
    0.25 ml
    0.25 ml
    0.25 ml
    0.25 ml

    Mix well , stand for 20 min at R.T
    2.5 ml
    2.5 ml
    2.5 ml
    2.5 ml
    2.5 ml
    2.5 ml

    Mix well , wait for 5 min then read at 505 nm aginst dist H2o

    Serum ALT and AST levels are increased in :
    • Acute hepatitis 
    • Exposure to drugs or toxic substances
    • Obstruction of bile ducts
    • Cirrhosis
    • Heart damage
    • liver tumers
    Serum ALT and AST levels are decreased in :
    • low levels of ALT and AST in blood are expected and normal in healthy individuals , healthy liver 
    Procedure with kits in lab :
    • Depend on kinetic technique
    • 500 µ AST or ALT reagent or (400µ R1 + 100µ R2),  incubate for 5 min at 37ͦ  C ,then add 50 µ  serum , Mix well and read .