Use of 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone chromogenic system in direct enzymic assay of uric acid in serum and urine.

1980 ◽  
Vol 26 (2) ◽  
pp. 227-231 ◽  
Author(s):  
P Fossati ◽  
L Prencipe ◽  
G Berti
Keyword(s):  
Uric Acid ◽  
Sulfonic Acid ◽  
Room Temperature ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Human Sera ◽  
Red Dye ◽  
Sample Measurement ◽  
Enzymic Assay ◽  
Colorimetric Procedure

Abstract A new direct colorimetric procedure for uric acid assay in serum or urine is described, utilizing a 3,5-dichloro-2-hydroxybenzene sulfonic acid/4-aminophenazone chromogenic system in the presence of horseradish peroxidase and uricase from Aspergillus flavus. This chromogen system has a high absorptivity, affording useful results with sample/reagent volume ratios as low as 0.025. The procedure is applicable to serum, plasma, or diluted urine. A single working reagent is used; the reaction is complete in less than 15 min at room temperature. The red dye formed is measured at 520 nm; a blank sample measurement is not needed. The standard curve for the method is linear for uric acid concentrations up to 1500 mumol/L. Average analytical recovery of uric acid in human sera and urine exceeded 99%; within-run and between-run precision studies showed CV's of less than or equal to 1.2 and less than or equal to 2.2%, respectively. The new procedure correlated well with the uricase/catalase and uricase/ultraviolet methods. The method is suitable for automation.

Use of 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone chromogenic system in direct enzymic assay of uric acid in serum and urine.

1980 ◽  
Vol 26 (2) ◽  
pp. 227-231 ◽  
Author(s):  
P Fossati ◽  
L Prencipe ◽  
G Berti
Keyword(s):  
Uric Acid ◽  
Sulfonic Acid ◽  
Room Temperature ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Human Sera ◽  
Red Dye ◽  
Sample Measurement ◽  
Enzymic Assay ◽  
Colorimetric Procedure

Abstract A new direct colorimetric procedure for uric acid assay in serum or urine is described, utilizing a 3,5-dichloro-2-hydroxybenzene sulfonic acid/4-aminophenazone chromogenic system in the presence of horseradish peroxidase and uricase from Aspergillus flavus. This chromogen system has a high absorptivity, affording useful results with sample/reagent volume ratios as low as 0.025. The procedure is applicable to serum, plasma, or diluted urine. A single working reagent is used; the reaction is complete in less than 15 min at room temperature. The red dye formed is measured at 520 nm; a blank sample measurement is not needed. The standard curve for the method is linear for uric acid concentrations up to 1500 mumol/L. Average analytical recovery of uric acid in human sera and urine exceeded 99%; within-run and between-run precision studies showed CV's of less than or equal to 1.2 and less than or equal to 2.2%, respectively. The new procedure correlated well with the uricase/catalase and uricase/ultraviolet methods. The method is suitable for automation.

Serum triglycerides determined colorimetrically with an enzyme that produces hydrogen peroxide.

1982 ◽  
Vol 28 (10) ◽  
pp. 2077-2080 ◽  
Author(s):  
P Fossati ◽  
L Prencipe
Keyword(s):  
Hydrogen Peroxide ◽  
Room Temperature ◽  
Volume Ratio ◽  
Glycerol Phosphate ◽  
Serum Triglycerides ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Human Sera ◽  
Sample Measurement ◽  
Colorimetric Procedure

Abstract In this direct colorimetric procedure, serum triglycerides are hydrolyzed by lipase, and the released glycerol is assayed in a reaction catalyzed by glycerol kinase and L-alpha-glycerol-phosphate oxidase in a system that generates hydrogen peroxide. The hydrogen peroxide is monitored in the presence of horseradish peroxidase with 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone as the chromogenic system. The high absorbance of this chromogen system at 510 nm affords useful results with a sample/reagent volume ratio as low as 1:150, and a blank sample measurement is not needed. A single, stable working reagent is used; the reaction is complete in 15 min at room temperature. The standard curve is linear for triglyceride concentrations as great as 13.6 mmol/L. Average analytical recovery of triglycerides in human sera is 100.1%, and within-run and between-run precision studies showed CVs of less than or equal to 1.6 and less than or equal to 3.0%, respectively. The method is suitable for automation.

Enzymic creatinine assay: a new colorimetric method based on hydrogen peroxide measurement.

1983 ◽  
Vol 29 (8) ◽  
pp. 1494-1496 ◽  
Author(s):  
P Fossati ◽  
L Prencipe ◽  
G Berti
Keyword(s):  
Hydrogen Peroxide ◽  
Room Temperature ◽  
Colorimetric Method ◽  
Creatinine Concentration ◽  
Standard Curve ◽  
Urine Creatinine ◽  
Analytical Recovery ◽  
Human Sera ◽  
Fuller’S Earth ◽  
Sample Measurement

Abstract We describe a new colorimetric method for measuring creatinine in serum and urine. Creatinine hydrolysis is catalyzed by creatinine amidohydrolase, and the creatine so produced is assayed in reactions catalyzed sequentially by creatine amidinohydrolase and sarcosine oxidase in a system that generates hydrogen peroxide. The hydrogen peroxide is measured at 510 nm in a reaction catalyzed by horseradish peroxidase, with 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone as the chromogen. This series of reactions is complete in 30 min at room temperature. A blank sample measurement corrects for endogenous creatine. The standard curve is linear for creatinine concentrations as great as 2.21 mmol/L. Analytical recovery of creatinine in human sera and urine averaged 99.8%. Within-run and between-run precision studies gave CVs of less than or equal to 3.3 and less than or equal to 4.3% for a serum with a creatinine concentration of 69 mumol/L. Results by this method agree well (r greater than 0.99) with those by both the enzymic ultraviolet method of Wahlefeld and the fuller's earth/Jaffé method.

Evaluation of a modified alcohol dehydrogenase assay for the determination of ethanol in blood.

1982 ◽  
Vol 28 (10) ◽  
pp. 2125-2127 ◽  
Author(s):  
A Poklis ◽  
M A Mackell
Keyword(s):  
Gas Chromatography ◽  
Reaction Time ◽  
Alcohol Dehydrogenase ◽  
Correlation Coefficients ◽  
Least Square ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Sigma Chemical ◽  
Enzymic Assay

Abstract We evaluated a new alcohol dehydrogenase (EC 1.1.1.1) enzymic assay (ADH-glycine, Sigma Chemical Co.) for the determination of ethanol in blood. This assay differs from the manufacturer's previous assay (ADH-pyrophosphate) in that glycine replaces pyrophosphate as the buffer and hydrazine replaces semicarbazide as the trapping agent. The standard curve for the assay was linear over blood ethanol concentrations of 0.50-5.00 g/L. The reaction time of the assay was 10 min. At 1.00 g/L within-run and between-run CVs were 3.96% (n = 20) and 4.01% (n = 20), respectively. Mean analytical recovery of ethanol added to whole blood at 0.50-5.00 g/L was 99.7% (SD 2.6%). We performed 100 consecutive clinical and forensic determinations by the ADH-glycine assay, the ADH-pyrophosphate assay, and gas chromatography. Correlation coefficients of the results by least-square linear regression were 0.995 for ADH-pyrophosphate vs ADH-glycine, and 0.990 for gas chromatography vs ADH-glycine. The major advantage of the ADH-glycine assay over the ADH-pyrophosphate assay is the shorter reaction time, 10 min vs 30 min.

Determination of uric acid on continuous-flow (AutoAnalyzer II and SMA) systems with a uricase/phenol/4-aminophenazone color test.

1978 ◽  
Vol 24 (2) ◽  
pp. 250-255 ◽  
Author(s):  
S Klose ◽  
M Stoltz ◽  
E Munz ◽  
R Portenhauser
Keyword(s):  
Uric Acid ◽  
Continuous Flow ◽  
Ascorbate Oxidase ◽  
Routine Test ◽  
Color Test ◽  
Clinical Laboratories ◽  
A Minor ◽  
Red Dye ◽  
Colorimetric Procedure

Abstract This report describes a new specific colorimetric procedure for uric acid assay with AutoAnalyzer II and SMA (Technicon) systems, made specific by the application of uricase. Hydrogen preroxide, formed in this reaction, effects the oxidative coupling of 4-aminophenazone and 2,4-dichlorophenol under the catalytic influence of peroxidase. The red dye formed is measured at 505 or 520 nm. A sample blank measurement is not necessary, and the reagents show very good stability. The test shows linearity up to 714 mumol of uric acid per liter. Results of thie method correlate very well with those by the uricase-ultraviolet and uricase--catalase methods. There is no interference by hemoglobin, bilirubin, lipemia, and various drugs, except a minor interference by alpha-methyldopa. Interference from ascorbate is eliminated by ascorbate oxidase. This method can be regarded as a considerably improved routine test for uric acid on continuous-flow systems in clinical laboratories as compared with the commonly used phosphotungstate method.

Semi-automated direct colorimetric measurement of creatine kinase isoenzyme MB activity after extraction from serum by use of a CK-MB-specific monoclonal antibody.

1988 ◽  
Vol 34 (3) ◽  
pp. 575-581 ◽  
Author(s):  
Y Landt ◽  
H C Vaidya ◽  
S E Porter ◽  
K Whalen ◽  
A McClellan ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Creatine Kinase ◽  
Room Temperature ◽  
Colorimetric Assay ◽  
Standard Curve ◽  
Colorimetric Measurement ◽  
Analytical Recovery ◽  
Colored Product ◽  
Quality Control Materials ◽  
Creatine Kinase Isoenzyme Mb

Abstract This semi-automated colorimetric assay for the MB isoenzyme of creatine kinase (EC 2.7.3.2) is based on a monoclonal antibody ("Conan-MB") specific for this isoenzyme and is a modification of a previously published method (Vaidya et al., Clin Chem 1986;32:657-63). A 0.64-cm bead coated with 2 to 3 micrograms of antibody is incubated with 100 microL of serum and 10 microL of 0.2 mol/L beta-mercaptoethanol for 1 h at room temperature, to extract CK-MB. The beads are washed with de-ionized water and incubated with CK substrate for 45 min at 37 degrees C. A solution containing trans-1,2-diaminocyclohexane-N,N,N', N'-tetraacetic acid, p-iodonitrotetrazolium violet, and diaphorase is added and the resulting colored product is measured at 492 nm. The standard curve is linear to 200 U of CK-MB per liter, and analytical recovery is 97-113%. Total assay CV for low (9.7 U/L) and high (50.7 U/L) quality-control materials was 14.1% (n = 1878) and 11.6% (n = 1842), respectively. CK-MB activity correlated well (r = 0.978, n = 226) with CK-MB measured by a two-site mass immunoassay, and 99.4% of samples with CK-MB greater than or equal to 12 U/L (n = 347) were verified by electrophoresis on agarose.

Enzymic assay of creatinine in serum and urine with creatinine iminohydrolase and glutamate dehydrogenase.

1982 ◽  
Vol 28 (7) ◽  
pp. 1461-1464 ◽  
Author(s):  
E Tanganelli ◽  
L Prencipe ◽  
D Bassi ◽  
S Cambiaghi ◽  
E Murador
Keyword(s):  
Glutamate Dehydrogenase ◽  
Reagent Blank ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Kinetic Test ◽  
Fuller’S Earth ◽  
Enzymic Assay ◽  
Serum And Urine

Abstract We describe an assay for creatinine in which it is converted by creatinine iminohydrolase (EC 3.5.4.21) into ammonia and N-methylhydantoin. The ammonia is subsequently assayed by use of alpha-ketoglutarate and glutamate dehydrogenase (EC 1.4.1.3). Use of NADPH as coenzyme eliminates all interferences from endogenous reactions. Endogenous ammonia in the sample is eliminated during a preincubation. The reaction reaches the endpoint in 15 min at working temperatures of 20-37 degrees C. No sample blank or reagent blank is needed. The standard curve is linear at least to 884 mumol (100 mg) of creatinine per liter. Average analytical recovery of creatinine in serum and urine is 99%. Within-run and between-run CVs are less than or equal to 2% and less than or equal to 6% for creatinine values of 335 mumol/L (38 mg/L) and 80 mumol/L (0 mg/L), respectively. Results by the described method (y) compare well with those by Jaffé's kinetic test (y = 1.01x -- 12.8), Berthelot/AutoAnalyzer method after treatment with immobilized creatinine iminohydrolase (y = 0.987x -- 13.2), Jaffé's test run on the SMA 12/60 (y = 1.011x -- 5.8), the Wahlefeld method (y = 1.014x -- 0.88), and Jaffé's test after deproteinization and absorption on fuller's earth (y = 0.985x -- 3.08). The method may be suitable for discrete, including centrifugal, automation.

New Chromogen for Assay of Glucose in Serum

1992 ◽  
Vol 38 (4) ◽  
pp. 522-525 ◽  
Author(s):  
R Reljic ◽  
M Ries ◽  
N Anić ◽  
B Ries
Keyword(s):  
Human Serum ◽  
Glucose Oxidase ◽  
Color Change ◽  
Colorimetric Assay ◽  
Volume Ratio ◽  
Glucose Determination ◽  
Analytical Recovery ◽  
Human Sera ◽  
Sample Measurement ◽  
Significant Interference

Abstract We describe a colorimetric assay for glucose determination in human serum, with use of the chromogen 2-amino-4-hydroxybenzenesulfonic acid (AHBS), glucose oxidase, and peroxidase. With this assay, glucose concentrations less than or equal to 27.8 mmol/L can be measured in serum, with a sample/reagent volume ratio as low as 0.0025. The chromogen itself is easily soluble in water and does not require other components for the color change, making the reagent composition less complex. A single working reagent is used, and the reaction is completed within 10 min at 37 degrees C. The absorbance of the yellow reaction product is measured at 415 nm, and a blank sample measurement is not needed. The average analytical recovery of glucose in different human sera was 97.6%, with no significant interference of reducing compounds in serum. The results of the recommended procedure correlated well with those of the phenol/4-aminoantipyrine method of Trinder.

Assay performance and clinical validation of a radioimmunoassay for serum thyroxine-binding globulin

2005 ◽  
Vol 24 (2) ◽  
pp. 115-119 ◽  
Author(s):  
Ivana Petrovic ◽  
Svetlana Savin ◽  
Dubravka Cvejic
Keyword(s):  
Specific Binding ◽  
Reference Ranges ◽  
Thyroxine Binding Globulin ◽  
Serum Thyroxine ◽  
Standard Curve ◽  
Coefficients Of Variation ◽  
Assay Performance ◽  
Analytical Recovery ◽  
Satisfactory Precision ◽  
Human Sera

A radioimmunoassay for the quantitation of serum thyroxine-binding globulin (TBG), RIA TBG (PEG) - INEP, was formed and characterised. Test sensitivity was found to be favorable (2.0 mg/L). The highly satisfactory precision of the assay was reflected in the low coefficients of variation (less than 5%) found for both intra- and interassay precision at three concentrations. Non-specific binding of the labeled antigen was less than 2% of the total binding. Linearity of the assay within the range of the standard curve (0-80.0 mg/L) was excellent (r =0.99) and the analytical recovery of TBG added to serum ranged from 91 to 115%. Reproducibility was checked by determining the concentration of TBG in samples of sera over a period of 32 days from the day of antigen iodination and F<Ftab 0.05, F0.05 7, 32 = 2.31. The assay was standardized for accuracy using Lyphochek Immunoassay Controls (Bio-Rad). A significant correlation was obtained between RIA TBG (PEG) and the Kodak Amerlite TBG Assay (r = 0.84; n = 25). The presented results show that RIA TBG (PEG) is a sensitive, precise and reliable method for determination of TBG in human sera. Assay reference ranges were established for sera from various subjects: euthyroid adults, hypothyroid and hyperthyroid patients and in healthy pregnant women.

The 4-hydroxybenzoate/4-aminophenazone chromogenic system used in the enzymic determination of serum cholesterol.

1978 ◽  
Vol 24 (12) ◽  
pp. 2161-2165 ◽  
Author(s):  
F Meiattini ◽  
L Prencipe ◽  
F Bardelli ◽  
G Giannini ◽  
P Tarli
Keyword(s):  
Ascorbic Acid ◽  
Uric Acid ◽  
Linear Regression ◽  
Serum Cholesterol ◽  
Reduced Glutathione ◽  
Cholesterol Oxidase ◽  
Dihydroxybenzoic Acid ◽  
Standard Curve ◽  
Analytical Recovery

Abstract A single reagent, containing cholesterol oxidase, cholesterol esterase, peroxidase, 4-hydroxybenzoate, and 4-aminophenazone, is used in determining serum cholesterol. Analysis time is 15 min, and the standard curve is linear to 6.0 g/liter. Analytical recovery of cholesterol was 100.1 +/- 0.4%. Within-run precision (CV) was less than or equal to 1.4 1.4%, between-run less than or equal to 4.8%. Comparison with results by a Liebermann Burchard method [Clin. Chim. Acta 5, 637 (1960)] gave a linear regression of y = 1.08x--0.05, with a correlation coefficient (r) of 0.985. Comparison with the Roeschlau enzymic method [J. Clin. Chem. Clin. Biochem, 12, 226 (1974)] gave y = 1.02x + 0.01 (r = 0.958). Comparison with the enzymic method of Allain et al. [Clin. Chem. 20, 470 (1974)] gave y = 1.01x--0.00 (r = 0.995). The following substances do not interfere up to the indicated concentrations (mg/liter): hemoglobin (5000), bilirubin (100), reduced glutathione (150), l-cysteine (400), urea (3000), creatinine (200), uric acid (200), d-glucose (10000), L-ascorbic acid (50), acetylsalicylic acid (500), L-DOPA (10), ergothioneine (1000), 2,5-dihydroxybenzoic acid (20), and 3,4-dihydroxybenzoic acid (10). Stored in an amber-colored bottle, the working reagent is stable for three months at 2--8 degrees C and for three weeks at 25 degrees C.

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