Estimation of alkaline phosphatase in serum
Introduction
- Phosphatase are enzymes which catalyze the splitting of phosphoric acid from certain mono phosphoric esters such as (glycerophosphates and phenyl phosphates)
- Alkaline phosphatase with a maximum activity at about PH 10
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Principle
- Alkaline phosphatase in serum acts on the substrate disodium phenyl phosphate buffered at PH 10 at 37ᵒ to liberate phenol
- This phenol condenses with 4- amino antipyrine in the presence of alkaline oxidizing agent potassium ferricyanide to give red colored complex which is measured at 520 nm
Normal range
- Alkaline phosphatase : 21 – 92 IU / L or 3 – 13 KA units
- Note : one IU of ALP activity = 7.13 KA units
Reagents
- Buffered substrate PH 10
- Phenol std
- Chromogen reagent 4 – amino antipyrine 0.6 % , potassium ferricyanide 2.4 %
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Procedure
Procedure
Reagent |
Blank |
Standard |
Test |
Buffered substrate PH 10 |
0.5 ml |
0.5 ml |
0.5 ml |
D / W |
1.55 ml |
1.55 ml |
1.55 ml |
Mix well and incubate at 37ᵒ for 15
min | |||
Serum |
– |
– |
50µ |
Phenol STD |
– |
50µ |
– |
Mix well and incubate at 37ᵒ for 15
min | |||
Colored reagent |
1 ml |
1 ml |
1 ml |
Mix well and read at 520 nm | |||
Calculation
- ALP activity (IU/L) =
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Serum ALP levels are increased in:
Serum ALP levels are increased in:
- Liver diseases: enzyme is produced by the hepatocytes canaliculi and hence
marked increase is seen in condition of obstruction of bile flow
- Extrahepatic biliary obstruction due to : stones in bile duct , cancer of head of pancreas
- Intrahepatic biliary obstruction due to : viral / drug induced hepatitis or malignancy
- Bone diseases: when osteoblastic activity in bone formation is take place
like in:
- Rickets : values of ALP more than 100 KA units can be seen
- Osteomalacia : less rise than in rickets
- Paget’s disease : very high values seen
- Others: fanconi’s syndrome, hyperparathyroidism
Serum ALP levels are decreased in:
- Severe anemia
- Malnutrition