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IVD

ALPHA AMYLASE

Direct Kinetic colorimetric method CNPG

FT En 08 Date of updating : 10/ 2020 Version B

IN VITRO DIAGNOSTIC USE

Manufacturer Use by In Vitro Diagnostic Temperature

Limitation

Catalogue number See insert Store away from light Sufcient

for < n > essais

Batch number

CINICAL SIGNIFICANCE

α-Amylases are hydrolytic enzymes which break down starch into maltose. In the human body

α-Amylases originate from various organs: pancreatic amylase is produced by the pancreas and

released into the intestinal tract; salivary amylase is synthesized in the salivary glands and secreted

into saliva. The amylase present in the blood is eliminated through the kidney and excreted into

the urine. Therefore, elevation of amylase activity in serum is reflected in a rise of urinary amylase

activity. Measurement of α–Amylase in serum and urine is mainly used for diagnosis of pancreatic

disorders as well as for detecting the development of complications.

PRINCIPLE

Kinetic colorimetric determination of the amylase activity in serum, plasma or urine depending on

the reaction:

5 CNPG

α–amylase

3 CNP + 2 CNPG

+ 2 G

+ 2 Glucose

α–Amylase hydrolyses 2-chloro-4-nitrophenyl-D-maltotrioside (CNPG

) to release 2-chloronitrophe-

nol and form 2-chloro-4-nitrophenyl-D-maltoside (CNPG

), maltriose (G

) and glucose (G). The rate

of formation of 2-chloro-nitrophenol measured by the variation in optical density at 405 nm per

minute is proportional to the amylase activity in the sample.

REAGENT COMPOSITION

Reagent CNPG

ready for use:

2-chloro-4-nitrophenyl-α–D-

maltotrioside (CNPG

) 2.5 mmol/l

Buffer MES pH = 6 50 mmol/l

NaCl 250 mmo/l

Ca (CH3 COO)

4.5 mmol/l

KSCN 375 mmol/l

Sodium Azide 152 mmol/l

REAGENT PREPARATION

- The reagent supplied is ready to use.

- Do not use the reagent if the OD of the reagent blank is greater than 0.500 at 405 nm

SAFETY CAUTIONS

Biomaghreb reagents are intended for use by qualified personnel for in vitro use (do not pipette by

mouth).

• Refer to the current SDS available on request or at www.biomaghreb.com;

• Verify the integrity of the reagents before use; and

• Disposal of waste: comply with the legislation in force.

For safety reasons, treat any specimen or reagent of biological origin as potentially infectious.

Respect the legislation in force.

SAMPLE PREPARATION

• Serum or plasma collected on heparin or iodoacetate (EDTA or citrate must not be used).

• Embarrassing haemolysis

• Urine

NOTE

The reagent should be stored at 15° - 25°C before use. DO NOT PIPE BY MOUTH to avoid contami-

nation with salivary amylase.

PRESERVATION AND STABILITY

• The reagent is stable at 2-8° until the expiry date, and for 30 days at room temperature

(20-25°C).

ADDITIONAL EQUIPMENT

• Basic equipment of the medical analysis laboratory;

• Spectrophotometer or Clinical Biochemistry Analyzer.

LINEARITY

• The reaction is linear up to 2000 U/L.

• If the ΔDO per min is > 0.630 repeat the determination by diluting the sample 1/10 with

NaCl 0.9 g/l.

PROCEDURE

Wavelength…………………………………….405 nm

Temperature…………………………………….37°C

Tank………………………………………….1 cm thick

Adjust the Zero of the spectrophotometer to air or distilled water.

In a tube introduce:

• α–amylase Reagent 1000 µl

Equilibrate to 37°C, add

• Sample Serum 25 µl or Urine 10 µl

Mix and incubate at 37°C for 1 minute.

Read the initial absorbance and start timer immediatety.

Read again at constant intervals for 3 minutes

CALCULATION

α–amylase (U/L) Sérum = ΔD.O / min / x 3178

α–amylase (U/L) Urine = ΔD.O / min / x 7829

REFERENCE VALUES

37°C

Serum, plasma

< 100 U/l

Urine < 380 U/l

REFERENCES

- Kaufman RA. Tietz NW-Clin Chem.26: 8461 (1980) ;

- Ranson JHC.Curr Prob Surg, 16:1 ( 1979) ;

- Young DS. et al, v. clin chem, 21 (1975).