Amylase

683

Enzymes

α-Amylase

Serum or heparinised plasma

Reagent: 3 x 50 mL,
Ready for use

Instructions for Use  |  Safety Data Sheet

For professional use only.

For many years, the levels of serum and plasma a-amylase in patients have provided needed evidence for the diagnosis of the acute pancreatitis (1,3). Early assay techniques were based on either a change in the absorption maxima of the complex between starch and iodine as the a-amylase degraded the starch; or a measurement of the increase in reducing groups as the starch was hydrolysed by the a-amylase (4). These methods are not as reliable and easy to quantitative as spectrophotometric methods using a defined substrate (5). Some methods are based on the production of NADH proportionate to the activity of the α-amylase. A defined substrate, such as maltotetraose, is degraded by a-amylase to produce glucose which can be measured in a coupled enzyme assay. However, this method necessitates the removal of endogeneous glucose which would give a high background to the assay (5). More recent methods are based on the production of p-nitro-phenol from defined oligosaccharide substrates with blocking groups attached on the terminal sugar. The action of the a-amylase on the oligosaccharides yields a variety of chain lengths after hydrolysis. These methods then use a variety of coupling enzymes to hydrolyse the resulting short chain oligosaccharides to produce p-nitropheno (5). The coupling enzymes contain residual a-amylase activity that may significantly reduce the stability of the reagent.

The α-Amylase assay involves the use of a chromogenic substrate, 2-chloro-4-nitrophenol linked with maltotriose (7). α-amylase hydrolyses the 2-chloro-4-nitrophenyl-α-D-maltotrioside (CNPG3) to release 2-chloro-aminophenol (CNP) and form 2-chloro-4-nitrophenyl- α-D-maltoside (CNPG2), maltotriose (G3) and glucose (G). The rate of formation of the 2-chloro-4-nitrophenol can be detected spectrophotometrically at 405 nm to give a direct measurement of α-amylase activity in the sample. The reaction is not readily inhibited by endogeneous factors

Store at 2 - 8°C and protect from direct light. When correctly stored, the reagents are stable up the expiry date reported on the label. A slight variation in the composition of the reagent may occur between batches, but this has no effect on the test results. After opening, the vials are stable 30 days if recapped immediately and protected from contamination, evaporation, direct light and stored at correct temperature.

Multicalibrator (R0300000006)
Normal control serum (R0400000006)
Pathological control serum (R0400000106)