00186 Rome (a) Tel. (+30) 06 #11 2316 Fax: (230) 06 4103079 Ema biBbolecnicas Webste: | Blotecnica Instruments § pA, Via Llcenza 18, Liquid Reagents - ready to use URIC ACID TBHBA Enzymatic, Colorimetric with ATCS* 2 Reagents Diagnostic reagent for quantitative in vitro determination (of uric acid in human serum, plasma or urine on photo- metric systems * navanced Turiiy Clearing Sytem; minimaes tubidty caused by lioemia [REF] (Com 422.8 x62.5mi 8x50_mi Reagent 1 8x125mi Reagent ‘Adéitionaly offered 41 6xSmi Control serum normal 4820 6x5mi Control serum pathologic 4830 3x3mi Mult calloraton serum ‘TEST PARAMETERS Method ‘Colorimetric, endpoint, increasing reaction, enzymatic Wavelength: 520 nm, Hg 846 nm (500-550 nm) Temperature: 20~25°C, 37°C Sample ‘Serum, heparin or EDTA plasma, urine Linear: {upto 20 mg/dL. (1190 umolL) Sensitivity: The lower limit of detection is 0.07 mg/dL (42 pmol), REAGENT COMPOSITION ‘COMPONENTS. ‘CONCENTRATION Reagent 1: Phosphate Buffer, pH 7.0 400. mmol TBHEA 1.25 mmol. Reagent 2: Phosphate Buffer, pH 7.0 100. mmol 4-Aminoantpytine 4.5 mmol. Ka (Fe(CN) 50 pmolt. POD. 210 KUL, Uricase 2150 UNL REAGENT PREPARATION Substrate Start ‘The reagents are ready to use, Sample Start: Mix 4 parts of Reagent 1 with t part of Reagent 2. (= Working Resgent) REAGENT STABILITY AND STORAGE Conditions: Protect from light. Close immediately after use. Do not freeze the reagents! ‘Avoid contamination Substrate Sta Stability: at2-8°C up to the expiration date Sample Start (Working Reagent): Stability, at2-8°C ‘3 months MATERIALS REQUIRED BUT NOT PROVIDED. NaCl solution (@ gL) General laboratory equipment INTERFERING SUBSTANCES no Interference upto: bilirubin Jo mgldt ‘iglyeerie 2000 mg/dl ‘hemoglobin 100 mg/dL ‘Ascorbic acd interferes even in minimal concentrations. For further information on interfering substances refer to Young DS (6). MANUAL TEST PROCEDURE Bring resgents and samples to room temperature. Substrate Start Pipette into test woes | Blonk | Si/Gal_| Sample Reagent 1 000 yl | 1000 yk 1000 pL ‘Sample or Sw/Cal = 20 pL [20 pL. Distilled water 20 2 Mx. incubate for 5 min. el 20-25°C377G.Then add Mx. Incubate for 30 min. at 20-25°C or for 10 min. at 37°. Measure absorbance of sample and sida. against reagent lank within 60 minutes Sample Start Pipette into test bes | Blank | Staal | Sample Working reagent 71000 wl [1000 ph [1000 pt ‘Sample or St6/Cal - 20 yt [20 yl Distilled water 20yL ‘Mix. Incubate for 30 min. at 20-25°C or for 10min. at 37°C. Measure absorbance of sample and std/eal. against reagent blank within 60 minutes. at15-25°C 2weeks Protect the Working Reagent from light! Note: It has to be mentioned, that the measurement is ‘not influenced by occasionally occuring colour changes, {a long as the absorbance of the Working Reagent is <0. at 546 nm, ‘SAMPLE PREPARATION Urine: Dilute urine 1+ 10 wih dst. water ‘SAMPLE STABILITY AND STORAGE Serumiplasma [3]: at20-25°C 3 days a 48°C days ‘at 20°C 6 months Urine (4 at20-25°C Aadays Freeze only once! Discard contaminated specimens. ‘CALCULATION ‘Serum/Plasma: ric Acid (mo/at) = 2A Sample I Uric Acie moldL] = -* SERBS x Conc. StslCal [mola] Urine: Uric Acid mp/at] = SA Semele Conc. Sid/Cal Imola} x 11 ‘A Std/Cal UNIT CONVERSION gla. x 59.48 = pmol. REFERENCE RANGE (1,5]* Serum/Plasm: Females Males mgt _[ moi |mgiat | mmo ‘Adu 26-60 | 195-957] 35-72 | 208-428 Females Males ‘Children. mel _| mmol [mpi [mmo 0-5 cay 49-78 [113-470] 19-79 | 13-470. =A yeors | 1.7=5.1 [101-303 2.2-5.7 [131340 B= 11 years | 3.0-64 [178-381 | 30-64 | 178-361 $2 14 years_[ 32-61 | 190-963] 32—7.4 | 190440) 15=17 years | 32-64 [190-361] 45-81 [268-482 rine: ‘assuming normal diet 800 mglbah (4.76 mmolBah) ‘assuming low purine diet | = 600 mg/24h 3.57 mmol24n) * Each laboratory should check the reference ranges are vanslerable to fs own pation population and determine own florence ranges necessay, aca ce nhac RO, ETEONC Aen et se pee sett Magnesspe Sees DIAGNOSTIC IMPLICATION [1,2] Uric acid and its salts are end products of the purine metabo- lem. In gout, the most common complication of hyperurce- mia, increased serum love's of uric acid lead to formation of monosodium urate crystals around the joints. Further causes Of elevated blood concentrations of uric acid are renal diseas- fe with decreased excration of waste products, starvation, drug abuse and increased alcohol consume as well as use of certain medicaments. High uric acid lavels also constitute an indirect risk factor for coronary heart disease. Hypouricemia is seldom observed and associated with rare hereditary metabol- ic disorders. ‘TEST PRINCIPLE Uric acid is oxidized to allantoin by urcase. The generated hydrogen peroxide reacts with 4-aminoantipyine and 2.4.6 ‘Wloromo-S-hydroxybenzoic acd (TBHBA) to quinoneimine. Une Acid +2 H:0 + 0; URICASE > atanton + C02 + H:O2 TBHBA + 4-AAP +H,0; POD > Quinoneimine +3 H:0 ABBREVATIONS: ~ ‘AAP = 4-Aminoantipytine POD = Peroxidase TBHBA = 2.4,6-Trbromo-3-Hydroxybenzoic Acid PERFORMANCE CHARACTERISTICS LINEARITY / MEASURING RANGE ‘The test has been developed to determine uric acid concen- trations within 8 measuring range from 0.07 — 20 mg/dl. (42 — 1190 pmolL). When values exceed this range, samples should be diuted 1+ 1 with NaCl soluton (9 gil) and reas- ‘sayed mulliplying the resuit by 2. PRECISION (at 37°C) Tna-assay,n=20 [Mean imgal] $0] Ova) imgidt) San 2S ‘004 Ts Sample 2 5.35 oo | 074 Sample 3 104 o0s_| 077 Tnterassay,n=20 [Mean Imola] SD) CVI imgidt ‘Sample f 2 0.04 1 ‘Sample 2 523 0.09 163 ‘Sample 3 8.98, ont 1.06 METHOD COMPARISON ‘A comparison between Blotecnica Uric acié TBHBA (y) and a ‘commercially avalable test (x) using 70 samples gave folow- ing results: y= 1.02 x 0.44 mg/l; r= 0.997. QUALITY CONTROL ‘All controls with Uric Acid values determined by this method ‘can be used. We recommend: 481 6xSmi_Controlserum normal 482 6x5mi_ Control serum pathologic. CALIBRATION ‘The assay requires the use of @ uric acid standard or calibra tor. We recommend: 483 -3x3mi Mut calbration serum CCalbratoc values have been made traceable to the reference ‘method gas chromatography‘sotope diuion mass specromety (GC1DMS), ‘AUTOMATION Applications for automated systems are available upon re- quest WARNINGS AND PRECAUTIONS 1. In very rare cases, samples of patients with gammopathy right give falsified results. 2. Please refer to the safety data sheets and take the neces- sary precautions forthe use of laboratory reagents. 3, For diagnostic purposes, the results should always be as- sesced with the patient's medical history, clinical examina- tions and ater findings, (00166 Rome (ta) ‘al (+39) 06 #11 2216 Fax (+20) 06 4102070 Erma tbotecnica.s Webste: wun bitenica.t J Bltecnica Instruments S pA, Vie Lcenza 18, WASTE MANAGEMENT lease refer to local requirements REFERENCES 1. Thomas L, Cincal Laborato Diagnostics. 1 ed. Frankur: TH ‘ocks Veiagspeselschat 1008p. 208-1 2. Newman Ds Pre CP. Renal urcton and ravogen metals. ‘Burts CA, Aehwoed ER ere. Tietz Tenbook of inal Chem- ty. 3* ea Phlaclpi: W-B Saunders Conpary, 1999p. 120470. 3. Gder WG, Zavia et a The Quay of Diagnostic Samples. ed Darmetac GIT Verag 2001 948-2 4. Guder WG, Zavla Be eta. The quality of Dapnoste Samples." fe Darmstast GT Verig: 2001p 523. 5. Newman JO, Pre PC. Real Kneien and nitogen metabolites. Burts CA, Ashwoed ER ears. Tet Textbook of Cincal Chemis- ty. 3" ed Phlacelpa: W.8 Saunders Company, 1900p. 1250 6, Young DS. Elects of Drage on Cina! Laboratory Tests 5 Volume 1 and 2 Washingen, DC: The Amerian Assia or (Cnial Chemisty Pres 200, C€ [vol 2" ot