H-1117 Budapest,

URIC ACID (URICASE/PAP)

Budafoki street 111-113 FREEZE DRIED
Tel.: +36-1-205-3617 Cat. No.: 40921 40962
15x20 ml 20X100 ml
Tel./Fax: +36-1-205-3616

Reagent kit for the quantitative determination of uric acid concentration in

serum and urine. Enzymatic colorimetric method. Uricase /PAP test. Calibration frequency
Two point calibration is recommended
In the human body uric acid is the end-product of purine metabolism. It is excreted - after reagent lot change,
by the kidney. Increases of uric acid in the serum plasma or urine can be due to the - as required following quality control procedures.
overproduction of purine containing molecules or to insufficient excretion. The
Calculation using calibration
concentration is increased in various renal diseases, with increased cell lysis in the
presence of tumors, leukemia, toxemia of pregnancy. Prolonged elevation of the Asample
concentration leads to gout. xC stan dard = C sample
Astandard
Principle
Uricase transforms Uric acid in the sample into Allantoin, Carbon dioxide (CO 2) and A = Absorbance, C = Concentration
Hydrogen peroxide (H2O2). By the action of Peroxidase (POD) and in the presence of
Quality control
phenol-derivative, DHBS and 4-Aminoantipyrine, Hydrogenperoxide gives a
A quality control program is recommended for all clinical laboratories. The analysis
coloured indicator reaction which can be measured at 520 nm. The increasing in
of control material in both the normal and abnormal ranges with each assay is
absorbance correlates with (is proportional to) the uric acid concentration of the
recommended for monitoring the performance of the procedure. Each laboratory
sample.
should establish corrective measures to be taken if values fall outside the limits.
Uric acid+ 2H2O + O2 Uricase
→ Allantoin+ CO2 + H2O2
PERFORMANCES DATA
2H2O2 + 4-Aminoantipyrine + DHBS POD
→ Red quinone+4H 2 O

The following data were obtained using the Hitachi 717 analyzer (37°C).
DHBS = 3,5 -Dichloro -2- hidroxy-benzenesulfonic acid
Reference values
Linearity
Female: 148-357 µmol/l (2,5-6,0 mg/dl)
The test is linear up to 1800 µmol/l (30,0 mg/dl).
Male: 200-416 µmol/l (3,4-7,0 mg/dl)
Urine: 250-750 mg/24 h
Sensitivity
It is recommended that each laboratory should assign its own normal range.
It is recommended that each laboratory establishes its own range of sensitivity as this
is limited by the sensitivity of the spectrophotometer used. Under manual conditions
Reagents
however, a change of 0.001 Abs is equivalent to 2.3 µmol/l (0,04mg/dl) Uric acid
1. Reagent (R1)
concentration at 492 nm.
Phosphate buffer, pH=7.5 50 mmol/l
DHBS 4 mmol/l
2. Reagent (R2) Precision
Uricase 80 U/l
4-Aminoantipyrine 1 mmol/l Reproducibility
Peroxidase 660 U/l Sample Average conc. (µmol/l) SD CV%
Sample I. 302 4.7 1.56
Samples Sample II. 707 8.4 1.19
Serum.
Urine diluted in ratio of 1:10 with distilled water. Repeatability
If the urine sample is opalic then incubate at 60°C for ten minutes. The ascorbic acid Sample Average conc. (µmol/l) SD CV%
in the urine sample interferes with the test, so use diluted sample.
Sample I. 230 4.07 1.77
PROCEDURE Sample II. 576 11.7 2.04

Preparation and stability of working reagent Correlation

Dissolve one vial of R2 in appropriate amount of Rl. Comparative studies were done to compare our reagent with our Uric acid ADPS
Stability: at: 20-25 °C: 7 days assay on 62 human serum samples.
at 2- 8 °C: 4 weeks The results from these studies are detailed below.
If the absorbance of working reagent is higher than 0.1 at 492 nm the reagent can not Correlation coefficient: r=0.9898
be used. Linear regression: y (µmol/l)= 0.959x+34.1
(x=Uric acid PAP reagent, y= Uric acid ADPS reagent).
Assay conditions
Wavelength: 520 (490-550) nm Specificity
Temperature: 37 °C No interfere with hemoglobin 18.6 µmol/l (120 mg/dl), bilirubin 855 µmol/l (50
Cuvette: 1 cm light path
mg/dl), lipid 1000 mg/dl, glucose 55.5 mmol/l (1000 mg/dl) and ascorbic acid 0.11
Read against: reagent blank
mmol/l (2 mg/dl).
Method: endpoint (increasing)
Note
Pipette into cuvette
Do not use reagents after the expiry date stated on each reagent container label. Do
not use products, test solutions and reagents described above for any purpose other
Blank Standard Sample than described herein.

Working reagent 1 ml 1 ml 1 ml For in vitro diagnostic use only.

Distilled water 20µl The following symbols are used on labels

Standard 20µl For in vitro diagnostic use

Sample 20µl Use by (last day of the month)

Mix and read the absorbance (A) after a 5-minute incubation at 37 °C. Temperature limitation

Calibration: (37°C, Uricase/PAP method) Batch Code

S1: Distilled water
S2: Uric acid standard Cat. No.: 50511 or Code
Roche C.F.A.S. (Calibrator for automated system)
Randox Calibration Serum Level I or Bibliography
Randox Calibration Serum Level II Barham, Trinder, Analys. 97, 142(1972)
Fossati Principe. Clin. Chem. 28, 227(1980)

Uric acid (Uricase/PAP), freeze dried Date of revision: 2009-12 09-EN-2009-07