OXALATE

COD 12539 1 x 25 mL COD 11839 5 x 25 mL

STORE AT 2-8ºC
Reagents for measurement of oxalate concentration OXALATE
Only for in vitro use in the clinical laboratory OXALATE OXIDASE/PEROXIDASE

PRINCIPLE OF THE METHOD

A25 A15
Oxalate in the sample generates, by means of the coupled reactions described below, a coloured complex that
can be measured by spectrophotometry 1,2. GENERAL Test name OXALATE OXALATE
Analysis mode Differential bir Differential bir
Oxalate oxidase Sample type URI URI
Oxalate + O2 CO2 + H2O2 Units mmol/L mmol/L
Peroxidase Reaction type increasing increasing
H2O2 + MBTH + DMAB Indamie Dye + H2O
Decimals 2 2
CONTENTS No. of replicates 1 1
COD 12539 COD 11839 Test name in patient report - -
1. Reagent 1 x 25 mL 5 x 25 mL PROCEDURE Reading Monochromatic Monochromatic
2. Tubes 1 x 20 5 x 20 Volumes Sample 13 13
A. Reagent 1 x 20 mL  Reagent 1 240 240
B1. Reagent 1 x 5 mL  Reagent 2 60 60
B2. Reagent 1 x 5 mL  Washing 1.2 1.2
S. Standard 1 x 5 mL  Predilution factor - -
Postdilution factor 2 2
COMPOSITION Filters Main 600 600
1. Reagent. Phosphate buffer 100 mmol/L, EDTA 5 mmol/L, preservatives, pH 7.0. Reference - -
WARNING: H317: May cause an allergic skin reaction. P302+P352: IF ON SKIN: Wash with plenty of Times Reading 1 75 s 72 s
soap and water. P333+P313: If skin irritation or rash occurs: Get medical advice/attention. Reading 2 450 s 456 s
2. Purifier tubes. Activated charcoal. Reagent 2 90 s 96 s
A. Reagent. Citric acid 100 mmol/L, 3-dimethylamino benzoic acid (DMAB) 0,25 mmol/L, 3-methyl-2- CALIBRATION Calibration type specific specific
benzothiazolinone hydrazone (MBTH) 0,1 mmol/L, pH 2.6. Calibrator replicates 3 3
B1. Reagent. Citric acid 100 mmol/L, preservatives, pH 5.6. Blank replicates 3 3
WARNING: H317: May cause an allergic skin reaction. P302+P352: IF ON SKIN: Wash with plenty of Calibration curve - -
soap and water. P333+P313: If skin irritation or rash occurs: Get medical advice/attention.
B2. Reagent. Oxalate oxidase > 300 U/L, peroxidase > 10 KU/L, after reconstitution. OPTIONS Blank absorbance limit 0.150 0.150
S. Oxalate standard. Oxalic acid 45 mg/L equivalent to 90 mg/L (1.0 mmol/L) of oxalate according to the Kinetic blank limit - -
dilution factor of the sample. Aqueous primary standard. Linearity limit 2 2
WARNING: H317: May cause an allergic skin reaction. P302+P352: IF ON SKIN: Wash with plenty of
soap and water. P333+P313: If skin irritation or rash occurs: Get medical advice/attention. REFERENCE VALUES
For further warnings and precautions, see the product safety data sheet (SDS). Urine3: 17.5-35.1 mg/24h = 0.20-0.60 mmol/24h.
STORAGE These ranges are given for orientation only; each laboratory should establish its own reference ranges.
Store at 2-8ºC.
Reagents are stable until the expiry date shown on the label when stored tightly closed and if contaminations are
QUALITY CONTROL
prevented during their use. It is recommended to use the Oxalate Control Urine (cod. 18062) to verify the performance of the measurement
Indications of deterioration: procedure. Oxalate concentrations are given on the vial label. Concentration value is traceable to the Oxalate
 Reagents: Presence of particulate material, turbidity, absorbance of the blank higher than 0.150 at 600 nm Standard. Traceability can be assured only if the BioSystems reagents and recommended measurement
(1 cm cuvette). procedure are used.
 Standard: Presence of particulate material, turbidity. Reconstitute the material with the volume of distilled water indicated in the label. Stable for 7 days at 2-8ºC.
Stable for 30 days at -20ºC (only freeze once).
REAGENT PREPARATION
Treat the Control in the analytical procedure as patient samples.
Reagents 1,2, A and the Standard are ready to use. The intervals of suggested acceptable values have been calculated from previous experience in interlaboratory
Reagent B: Reconstitute the Reagent B2 with the contents of the Reagent B1 vial. Mix gently. Stable for 60 days variability and are given for orientation only; each laboratory should establish its own precision.
at 2-8ºC.
Reagents open and kept in the refrigerated compartment of the analyzer are stable 60 days. METROLOGICAL CHARACTERISTICS
ADDITIONAL EQUIPMENT The following data were obtained using an A25 analyser. Results are similar with A15. Details on evaluation data
are available on request.
 Analyzer, spectrophotometer or photometer able to read at 600  20 nm.
 Linearity limit: 180 mg/L = 2 mmol/L. For higher values dilute pretreated sample 1/2 with distilled water and
SAMPLES repeat measurement.
Urine: Collect a 24-hour urine specimen using HCI as a preservative. It is recommended that patients refrain  Detection limit: 1.8 mg/L = 0.02 mmol/L.
from taking vitamin C rich food for at least 48 hours prior to urine collection.
Oxalate in acidified urine is stable for 7 days at 2-8ºC.  Repeatability (within run):

PROCEDURE Mean oxalate concentration CV n

Sample pretreatment 28.8 mg/L = 0.32 mmol/L 1.0 % 20

The standard do not require pretreatment. 109 mg/L = 1.21 mmol/L 0.4 % 20

1. Pipette into a test tube:  Reproducibility (run to run):

Urine 1 mL Mean oxalate concentration CV n
Reagent 1 1 mL 28.8 mg/L = 0.32 mmol/L 3.7 % 25
2. Shake thoroughly. Check the pH. It should be between 5 and 7 pH, if not adjust the pH with hydrochloric acid 109 mg/L = 1.21 mmol/L 1.5 % 25
1 mol/L or sodium hidroxide 1 mol/L.
 Trueness: Results obtained with this procedure did not show systematic differences when compared with a
3. Pour the diluted sample in a purifier tube. Mix for 5 minuts by intermittent mixing with a mixer.
reference procedure. Details of the comparison experiments are available on request.
4. Centrifuge the tubs for 10 minuts at 3000 rpm.
 Interferences: Bilirubin (< 30 mg/dL) and hemolysis (<450 mg/dL) do not interfere. Ascorbic acid (> 16
5. Determine the oxalate concentration in the supernatant, it is estable 7 days at 2-8ºC. mmol/L) may affect the results. Other drugs and substances may cause interference 4.
Manual procedure (Note 1)
DIAGNOSTIC CHARACTERISTICS
1. Bring the Reagents and the instrument to 37ºC. Oxalate is an end product of metabolism, predominantly derived from the degradation of glyoxylate and glycine.
2. Pipette into a cuvette: It is eliminated entirely urine and only about 15% of urinary oxalate is derived directly from dietary sources.
Reagent Blank Sample/Standard Hyperoxaluria is a powerful promoter of calcium oxalate stone formation. An increased excretion of oxalate in
urine may occur as a result of an excessive ingestion of oxalate rich foods, because of malabsorption due to
Distilled water 45 µL  different gastrointestinal disorders (enteric hyperoxaluria) or because of an inborn error of metabolism (primary
Reagent A 800 µL 800 µL hyperoxaluria)3. Low oxalate values in urine are associated with hyperglycinemia or hyperglycinuria.
Sample/Oxalate Standard (S)  45 µL Clinical diagnosis should not be made on the findings of a single test result, but should integrate both clinical and
3. Mix and insert the cuvette into the instrument. Start the stopwatch. After 5 minutes, read the absorbance (A 1) laboratory data.
at 600 nm. NOTES
4. Pipette into a cuvette:
1. The quantity of Oxalate excreted during 24 hour period can be calculated multiplying the concentration value
Reagent B 200 L 200 L by the total volume of urine voided.
2. This reagent may be used in several automatic analysers. Instructions for many of them are available on
5. Mix and insert the cuvette into the instrument. Start the stopwatch. After 5 minutes, read the absorbance (A2) request.
at 600 nm.
6. The Oxalate concentration in the sample is calculated using the following general formula: BIBLIOGRAPHY
1. Laker M.F, Hofmann A.F, Meeuse B.J.D. Spectrophotometric determination of urinary oxalate with oxalate
(A2- 0.81 x A1) Urine - (A2- 0.81 x A1) Blank x 90 mg/L oxalate oxidase prepared from moss. Clin Chem 1980; 26:827-830.
2. Ngo T.T, Lenhoff H.M. A sensitive and versatile chromogenic assay for peroxidase and peroxidase-coupled
(A2- 0.81 x A1) Standard - (A2- 0.81 x A1) Blank x 1.0 mmol/L oxalate
reactions. Analytical Biochem 1980; 105:389-397.
3. Tietz Textbook of Clinical Chemistry and Molecular Diagnostics, 4th ed. Burtis CA, Ashwood ER, Bruns DE.
Automated procedure (Note 1,2) WB Saunders Co, 2005.
It is recommended to do a reagent blank every day and a calibration at least every 60 days, after reagent lot 4. Young DS. Effects of drugs on clinical laboratory tests, 5th ed. AACC Press, 2000.
change or as required by quality control procedures.

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