Biosystems Alkaline Phosphatase
PRINCIPLE OF THE METHOD
- Alkaline phosphatase (ALP) catalyzes in alkaline medium the transfer of the phosphate group from 4-nitrophenylphosphate to 2-amino-2-methyl-1-propanol (AMP), liberating 4-nitrophenol.
- The catalytic concentration is determined from the rate of 4-nitrophenol formation, measured at405 nm1
- A. Reagent: 2-Amino-2-methyl-1-propanol 0.4 mol/L, zinc sulfate 1.2 mmol/L,
- N-hydroxyethylethylenediaminetriacetic acid 2.5 mmol/L, magnesium acetate 2.5 mmol/L, pH10.4.
- B. Reagent: 4-Nitrophenylphosphate 60 mmol/L.
- Store at 2-8ºC.
- Reagents are stable until the expiry date shown on the label when stored tightly closed and if contaminations are prevented during their use.
- Indications of deterioration: Reagents: Presence of particulate material, turbidity, absorbance of the blank over 1.200 at405 nm (1 cm cuvette).
- Working Reagent:• Cod. 11592 and 11593: Transfer the contents of one Reagent B vial into a Reagent A bottle.
- Mix gently. Other volumes can be prepared in the proportion: 4 mL Reagent A + 1 mL Reagent B. Stable for 2 months at 2-8ºC.
Cod. 11598: Transfer 25 mL of one Reagent B vial into a Reagent A bottle. Mix gently.
- Other volumes can be prepared in the proportion: 4 mL Reagent A + 1 mL Reagent B. Stable for 2 months at 2-8ºC.
- Analyzer, spectrophotometer or photometer with cell holder thermostatable at 25, 30 or 37ºC and able to read at 405 nm.
Cuvettes with 1 cm light path.
- Serum and plasma collected by standard procedures.
- Alkaline phosphatase in serum or plasma is stable for 7 days at 2-8ºC. Heparin may be used as anticoagulant.
- Alkaline phosphatase catalyzes the hydrolysis of organic phosphate monoesters at alkaline pH.
- The enzyme is present in practically all tissues of the body, especially at or in the cell membranes, and it occurs at particularly high concentrations in placenta, intestinal epithelium,
kidney tubules, osteoblasts and liver.
- The form present in the sera of normal adults originates mainly in the liver and bone.
- Elevated serum ALP is found in patients with bone disease associated with increased osteoblastic activity (Paget’s disease, primary and secondary hyperparathyroidism, bone
tumors, rickets, osteomalacia, bone fractures) and also in patients with hepatobiliary disease(obstructive jaundice, hepatitis, hepatotoxicity caused by drugs, liver cancer).
- Physiological changes, such as bone growth and pregnancy, may cause increases in ALP levels4,5
- Clinical diagnosis should not be made on the findings of a single test result, but should integrate both clinical and laboratory data.