COD 11588 COD 11589 CHOLINESTERASE (CHE)

1 x 50 mL 4 x 50 mL
STORE AT 2-8ºC
Reagents for measurement of cholinesterase concentration CHOLINESTERASE (CHE)
Only for in vitro use in the clinical laboratory BUTYRYLTHIOCHOLINE

PRINCIPLE OF THE METHOD REFERENCE VALUES

Cholinesterase (CHE) catalyzes the hydrolysis of butyrylthiocholine to thiocholine and butyric acid. The catalytic Cholinesterase (37ºC):
concentration is determined from the rate of decrease of hexacyanoferrate (III), measured at 405 nm, by means
of the following reactions1,2,3. Men 4620-11500 U/L = 76.9-191 µkat/L
CHE Women 3930-10800 U/L = 65.5-180 µkat/L
Butyrylthiocholine + H2O Thiocholine + Butyric acid
Dibucaine number (%)2:
2 Thiocholine + 2 OH- + 2 Hexacyanoferrate (III) Dithiobis(choline) + 2 Hexacyanoferrate (II) + H2O
Normal homozygotes: > 75%
The atypical isoenzyme of cholinesterase (AA) can be estimated through the “dibucaine number” indicating the Intermediate heterozygotes: 45-72 %
inhibition of enzyme activity in the presence of dibucaine and expressed in per cent2,3. Atypical homozygotes: < 30 %

These ranges are given for orientation only; each laboratory should establish its own reference ranges.
CONTENTS
COD 11588 COD 11589 QUALITY CONTROL
A. Reagent 1 x 40 mL 4 x 40 mL Each laboratory should establish its own internal Quality Control scheme and procedures for corrective action if
B. Reagent 1 x 10 mL 4 x 10 mL controls do not recover within the acceptable tolerances.

COMPOSITION METROLOGICAL CHARACTERISTICS

A. Reagent. Pyrophosphate 95 mmol/L, Hexacyanoferrate (III) 2.5 mmol/L, pH 7.6. − Detection limit: 123 U/L = 2.05 µkat/L.
B. Reagent B. Butyrylthiocholine 60 mmol/L. − Linearity limit: 25000 U/L = 417 µkat/L. For higher values dilute sample 1/2 with distilled water and repeat
measurement.
STORAGE − Repeatibility (within run):
Store at 2-8ºC. Mean Concentration CV n
Reagents are stable until the expiry date shown on the label when stored tightly closed and if contaminations
are prevented during their use. 5416 U/L = 90.27 µkat/L 1.0 % 20
Indications of deterioration: 11279 U/L = 188.0 µkat/L 0.7 % 20
Reagents: Presence of particulate material, turbidity, absorbance of the blank below 1.300 at 405 nm (1 cm
cuvette). − Reproducibility (run to run):
Mean Concentration CV n
AUXILIARY REAGENTS 1.0 % 25
5416 U/L = 90.27 µkat/L
C. Reagent (cod 11578): Dibucaine 0.3 mmol/L, after reconstitution. 10 x 15 mL. 0.6 % 25
11279 U/L = 188.0 µkat/L
REAGENT PREPARATION − Sensitivity: 0.015 ∆mA⋅L/U⋅min = 0.91 ∆mA ⋅L/µkat⋅min.
Working Reagent for total cholinesterase: Pour the contents of the Reagent B into the Reagent A bottle. Mix − Trueness: Results obtained with this reagent did not show systematic differences when compared with
gently. Other volumes can be prepared in the proportion: 4 mL Reagent A + 1 mL Reagent B. Stable for 3 days reference reagents. Details of the comparison experiments are available on request.
at 2-8ºC.
Working Reagent with dibucaine (Note 1): reconstitute the contents of a Reagent C vial with 15 mL of the − Interferences: Lipemia (triglycerides < 10 g/L) and bilirubin (< 20 mg/dL) do not interfere. Hemolysis may affec
Working Reagent for total cholinesterase. Swirl gently. Stable for 3 days at 2-8ºC. the results. Other drugs and substances may interfere5.
These metrological characteristics have been obtained using an analyzer. Results may vary if a different
ADDITIONAL EQUIPMENT instrument or a manual procedure are used.
− Analyzer, spectrophotometer or photometer with cell holder thermostatable at 37ºC and able to read at 405
nm. DIAGNOSTIC CHARACTERISTICS
− Cuvettes with 1 cm light path. Serum cholinesterase is also called pseudocholinesterase to differentiate it from the cholinesterase present in
erythrocytes and in nerve endings. Cholinesterase is synthesized in the liver, and it can be used as a test of liver
SAMPLES1 function. A decrease in its activity reflects impaired synthesis.
Serum or plasma collected by standard procedures. Heparin and EDTA are recommended as anticoagulants. Its measurent is of particular value in the diagnostic of patients with atypical forms of the enzyme and in
Cholinesterase in serum or plasma is stable for 14 days at 2-8ºC. organophosphorus insecticide poisoning. Patients with the atypical form of the enzyme presents high sensitivity
towards suxamethonium, a drug used as muscle relaxant in surgery. Its identification by means of dibucaine
number is important to prevent the prolonged apnea caused by the administration of this drug4,6.
PROCEDURE
Changes in serum cholinesterase can also occur in other conditions. It is decreased in acute infections,
1. Bring the Working Reagent and the instrument to reaction temperature.
pulmonary embolism, muscular dystrophy and myocardial infarction4,6.
2. Pipette into a cuvette (Note 2):
Clinical diagnosis should not be made on the findings of a single test result, but should integrate both clinical
Working Reagent (Note 3) 1.5 mL and laboratory data.
Sample 25 µL
NOTES
3. Mix and insert the cuvette into the photometer. Start the stopwatch. 1. The working reagent with dibucaine is used only to determine the dibucaine number.
4. After 90 seconds, record initial absorbance and at 30 second intervals thereafter for 90 seconds. 2. These reagents may be used in several automatic analysers. Instructions for many of them are available on
5. Calculate the difference between consecutive absorbances, and the average absorbance difference per request.
minute (∆A/min). 3. Use working reagent without dibucaine to measure total cholinesterase and working reagent with dibucaine
to measure the non inhibited cholinesterase.
CALCULATIONS
The cholinesterase concentration in the sample is calculated using the following general formula: BIBLIOGRAPHY
1. DGKC. Proposal of standard methods for the determination of enzyme catalytic concentrations in serum and
6 plasma at 37ºC. II Cholinesterase. Eur J Clin Chem Chim Biochem 1992; 30: 163-170.
Vt × 10
∆A/min × = U/L 2. Panteghini M and Bonora R. Evaluation of a new continuous colorimetric method for determination of serum
ε × l × Vs
pseudo-cholinesterase catalytic activity and its application to a centrifugal fast analyser. J Clin Chem Clin
The molar absorbance (ε) of the dye at 405 nm is 927, the lightpath (l) is 1 cm, the total reaction volume (Vt) is Biochem 1984; 22: 671-676.
1.525, the sample volume (Vs) is 0.025, and 1 U/L are 0.0166 µkat/L. The following formulas are deduced for 3. Whittaker M, Britten JJ and Dawson PJ. Comparison of a commercially available assay system with two
the calculation of the catalytic concentration: reference methods for the determination of plasma cholinesterase variants. Clin Chem 1983; 29: 1746-1751.
4. Tietz Textbook of Clinical Chemistry, 2nd edition. Burtis CA, Ashwood ER. WB Saunders Co., 1994.
x 65804 = U/L
∆A/min 5. Young DS. Effects of drugs on clinical laboratory tests, 3th ed. AACC Press, 1997.
x 1097 = µkat/L
6. Friedman and Young. Effects of disease on clinical laboratory tests, 3th ed. AACC Press, 1997.
Dibucaine number (%) = 100-(100 x CHE non inhibited / CHE total)

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