New saccharogenic determination of alpha-amylase in serum and urine.

1979 ◽  
Vol 25 (2) ◽  
pp. 215-217 ◽  
Author(s):  
L van Leeuwen
Keyword(s):  
Human Serum ◽  
Amylase Activity ◽  
Glucose Dehydrogenase ◽  
Alpha Amylase ◽  
Endogenous Glucose ◽  
Alpha Glucosidase ◽  
Serum And Urine

Abstract I describe a new kinetic enzymatic saccharogenic method for assaying alpha-amylase in human serum and urine. alpha-Amylase liberates maltose from starch. This is successively acted on by alpha-glucosidase, mutarotase, and glucose dehydrogenase. The resulting conversion of NAD+ to NADH, measured at 340 nm, during a 20-min incubation reflects amylase activity. Endogenous glucose is destroyed before measurement of amylase activity is begun.

Enzyme-coupled ultraviolet determination of alpha-amylase activity with the GEMSAEC centrifugal analyzer.

1978 ◽  
Vol 24 (9) ◽  
pp. 1620-1624 ◽  
Author(s):  
W H Porter ◽  
R E Roberts
Keyword(s):  
Linear Response ◽  
Amylase Activity ◽  
Kinetic Studies ◽  
Alpha Amylase ◽  
Coefficients Of Variation ◽  
Endogenous Glucose ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Alpha Glucosidase ◽  
As Effects

Abstract We evaluated the Harleco alpha-glucosidase/hexokinase/glucose-6-phosphate dehydrogenase-coupled alpha-amylase method, bu use of the GEMSAEC centrifugal analyzer. Performance evaluation included kinetic studies of substrate and maltose hydrolysis as well as effects of endogenous glucose and fructose. The reagent was found to give a linear response with alpha-amylase activity to greater than 1200 U/liter. Within-run precision resulted in coefficients of variation (CV) of 0.9 to 3.2% over the range studied. Day-to-day precision corresponded to CV's of 2.4 to 4.4% over the same range of alpha-amylase procedure was found to be good (r = 0.997) for patients' sera examined.

Isopropylidine maltoheptosyl fructofuranoside, doubly blocked substrate for determination of endoamylase activity

1991 ◽  
Vol 37 (8) ◽  
pp. 1323-1328
Author(s):  
Z Ogawa ◽  
Y Matsubayashi ◽  
S Satoh ◽  
N Orita ◽  
H Itoh
Keyword(s):  
Human Serum ◽  
Coefficient Of Variation ◽  
Oxidation Rate ◽  
Amylase Activity ◽  
Mannitol Dehydrogenase ◽  
Coupled Reactions ◽  
Alpha Glucosidase ◽  
Serum And Urine

Abstract We synthesized o-(4,6-o-isopropylidene-alpha-D-glucopyranosyl)-(1----4)- [o-alpha-D-glucopyranosyl-(1----4])5-o-alpha-D-glucopyranosyl-(1----2)- alpha-D-fructofuranoside (IPG7F) and developed an assay for determining the activity of amylase in human serum and urine by using this substrate. Glucoamylase, alpha-glucosidase, and mannitol dehydrogenase are used as coupling enzymes. The coupled reactions are monitored by continuously measuring the oxidation rate of NADH. In this procedure, various substances in the test specimens do not interfere with the detection of amylase activity. Exactly one molecule of NADH is oxidized by one attack of amylase on the substrate, although four products can be produced in the reaction. The within-assay coefficient of variation (CV) ranged from 1.0% to 4.1% and the between-assay CV ranged from 2.6% to 5.3%. The results of our new assay correlate well with those of the amylase assay involving p-nitrophenol maltoheptaoside as substrate (r = 0.978) and with those of the amylase assay involving maltopentaose (r = 0.987).

Determination of alpha-amylase activity: methods comparison and commutability study of several control materials

1995 ◽  
Vol 41 (3) ◽  
pp. 435-438 ◽  
Author(s):  
G Gubern ◽  
F Canalias ◽  
F J Gella
Keyword(s):  
Human Serum ◽  
Amylase Activity ◽  
Alpha Amylase ◽  
Human Origin ◽  
Serum Samples ◽  
Chromophore Group ◽  
Laboratory Control ◽  
Wide Range ◽  
Human Sera

Abstract Six different methods for alpha-amylase determination were compared by assaying human serum samples covering a wide range of alpha-amylase values. All the methods studied use as substrate a maltooligosaccharide with a chromophore group at the reducing end; some are chemically blocked at the nonreducing end. Intermethod comparison by regression and correspondence analyses showed significant differences for two methods. The commutability of 12 commercial control materials containing alpha-amylase was also assessed by the different methods in comparison with human serum specimens containing the pancreatic and salivary isoenzymes. We also studied the behavior of pancreatic and salivary materials prepared in our laboratory. Control materials with alpha-amylase of non-human origin were not commutable with the enzyme in human sera and should not be used for intermethod calibration.

Specific immunoassay of alpha-amylase isoenzymes in human serum.

1985 ◽  
Vol 31 (8) ◽  
pp. 1331-1334 ◽  
Author(s):  
M Gerber ◽  
K Naujoks ◽  
H Lenz ◽  
W Gerhardt ◽  
K Wulff
Keyword(s):  
Monoclonal Antibody ◽  
Human Serum ◽  
Enzyme Immunoassay ◽  
Amylase Activity ◽  
Alpha Amylase ◽  
Pancreatic Amylase ◽  
Routine Method ◽  
Salivary Amylase ◽  
Amylase Isoenzymes

Abstract A monoclonal antibody (66C7) was prepared that specifically binds human salivary amylase (EC 3.2.1.1); it cross reacts with human pancreatic amylase by less than 1%. Two procedures are described for determination of isoamylases in human serum with this antibody: an enzyme immunoassay for determining amylase of salivary origin, and a routine method in which this amylase is immunoprecipitated and the remaining (pancreatic) amylase activity is assayed. Results by the two methods correlate well.

Rapid determination of alpha-amylase activity by use of a new chromogenic substrate.

1987 ◽  
Vol 33 (4) ◽  
pp. 524-528 ◽  
Author(s):  
G Dupuy ◽  
G Hilaire ◽  
C Aubry
Keyword(s):  
Amylase Activity ◽  
Rapid Determination ◽  
Molar Absorptivity ◽  
Cleavage Product ◽  
Alpha Amylase ◽  
Lag Phase ◽  
Chromogenic Substrate ◽  
Salivary Alpha Amylase ◽  
Alpha Glucosidase

Abstract A new chromogenic substrate that is blocked at the nonreducing end, 4,6-benzylidene-alpha-D-4-nitrophenylmaltoheptaoside, is used to determine alpha-amylase (EC 3.2.1.1) activity in serum in a coupled assay with alpha-glucosidase (EC 3.2.1.20) and glucoamylase (EC 3.2.1.3) as auxiliary enzymes. The duration of the lag phase between 25 and 37 degrees C is less than 90 s, and the molar absorptivity of 4-nitrophenol is constant. The main cleavage product of the substrate by human pancreatic and salivary alpha-amylase is 4-nitrophenylmaltoside; in the presence of the auxiliary enzymes, greater than 95% of hydrolyzed substrate is accounted for as 4-nitrophenol. The combined reagent is stable for at least 20 days at 2-8 degrees C; precision is good, with CVs ranging from 1.7 to 3.3%; and the correlation of results with those by the 4-nitrophenylmaltoheptaoside method is excellent. Heparin (40 kilo-int. units/L), ascorbic acid (2.8 mmol/L), bilirubin (430 mumol/L), hemoglobin (170 mumol/L), glucose (55 mmol/L), and triglycerides (11 mmol/L) do not interfere in the assay.

Electrophoretic pattern of amylase isoenzymes in serum and urine of normal persons.

1976 ◽  
Vol 22 (4) ◽  
pp. 439-444 ◽  
Author(s):  
M Otsuki ◽  
S Saeki ◽  
H Yuu ◽  
M Maeda ◽  
S Baba
Keyword(s):  
Thin Layer ◽  
Human Serum ◽  
Salivary Glands ◽  
Clinical Medicine ◽  
Amylase Activity ◽  
Electrophoretic Pattern ◽  
Normal Pattern ◽  
Amylase Isoenzymes ◽  
Serum And Urine

Abstract We separated and measured amylase isoenzymes in the serum and urine of 3036 normal persons by electrophoresis on a thin layer of polyacrylamide gel. We wished to establish the normal pattern of these isoenzymes and to evaluate the usefulness of this method of electrophoresis in clinical diagnosis. Results for patients with hyper- or hypofunctioning pancreas and salivary glands suggested that essentially all the isoamylases in human serum and urine are derived from the salivary glands and the pancreas, and revealed that isoamylases of more than 98% of normal persons consisted of two major isoenzymes and two to three minor ones. Although these observations indicate that data on changes in the proportion of amylase activity of each isoenzyme can be useful in clinical medicine, the following points should be remembered: (a) quantitative differences in the isoenzyme pattern were observed, depending upon the condition of the samples; (b) because the proportion of isoamylase activity in serum of different normal persons differs, seriatim determination of amylase isoenzymes is necessary; and (c) because five different genetically controlled types of isoamylases were observed in normal persons, genetic investigations are also necessary.

Fluorescence depolarization assay for quantifying alpha-amylase in serum and urine.

1985 ◽  
Vol 31 (9) ◽  
pp. 1478-1480 ◽  
Author(s):  
M Hofman ◽  
M Shaffar
Keyword(s):  
Human Serum ◽  
Calibration Curve ◽  
Fluorescence Polarization ◽  
Alpha Amylase ◽  
Pancreatic Amylase ◽  
Liquid Form ◽  
Fluorescence Depolarization ◽  
Endogenous Glucose ◽  
Hydrolysis Of ◽  
Serum And Urine

Abstract We have developed a new method for quantifying alpha-amylase (EC 3.2.1.1) in serum and urine by fluorescence depolarization. Amylase in the sample catalyzes the hydrolysis of the substrate, a fluorescein-labeled amylose. This results in decreased fluorescence polarization, owing to the increased rate of rotation of the amylose fragment relative to the intact substrate. The TDx amylase assay is calibrated with six human-serum-based pancreatic amylase calibrators. Amylase activities are determined by interpolation from the calibration curve, which is stored in the TDx analyzer's memory. Results correlate well with those by the Du Pont aca assay and the Beckman "DRI-STAT" assay. Endogenous glucose does not interfere. CVs are less than 6%, and the reagents are stable in liquid form.

Suitability of control materials for determination of alpha-amylase activity.

1981 ◽  
Vol 27 (6) ◽  
pp. 806-815 ◽  
Author(s):  
J P Bretaudiere ◽  
R Rej ◽  
P Drake ◽  
A Vassault ◽  
M Bailly
Keyword(s):  
Quality Control ◽  
Human Serum ◽  
Amylase Activity ◽  
Linear Representation ◽  
Alpha Amylase ◽  
Reference Groups ◽  
The Matrix ◽  
Components Analysis ◽  
Pancreatic Pathology

Abstract The suitability of control materials for determination of alpha-amylase activity was assessed in comparison with reference groups of authentic human serum specimens containing alpha-amylase of either pancreatic or salivary origin, specimens from patients with no pancreatic pathology, and normal specimens to which porcine pancreatic alpha-amylase was added. After determination of alpha-amylase activity by 11 commonly used techniques (five different principles), the results were processed by both classical (linear representation, regression) and multivariate (correspondence analysis, principal-components analysis) statistical techniques. Specimens containing porcine pancreatic alpha-amylase did not behave like any of the other groups. We conclude that porcine enzyme should not be used for interlaboratory quality-control surveys or intermethod comparison studies. Determination of human salivary and pancreatic alpha-amylase showed intermethod biases similar to those for authentic patients' specimens. Human salivary alpha-amylase, both because of its behavior and its commercial availability, is a satisfactory source for alpha-amylase activity of quality-control specimens. The nature of the matrix (polyvinylpyrrolidone, albumin, delipidated serum, bovine serum, or human serum) little influenced the behavior of the specimens for any of the methods studied.

New, Simple Maltogenic Assay for Mechanized Determination of Alpha-Amylase Activity in Serum and Urine

1975 ◽  
Vol 21 (6) ◽  
pp. 694-702 ◽  
Author(s):  
Henning F Proelss ◽  
Billy W Wright
Keyword(s):  
Continuous Flow ◽  
Flow System ◽  
Amylase Activity ◽  
Alpha Amylase ◽  
Automated System ◽  
Normal Range ◽  
Clin Pathol ◽  
Reducing Substances ◽  
Serum And Urine

Abstract We report a new method for the mechanized determination of serum and urinary alpha-amylase by use of a continuous-flow system, based on the measurement of maltose formed by incubating the sample with amylodextrin at pH 7 and 40 °C. After dialysis, maltose is converted enzymatically to glucose, which is measured by Trinder's glucose oxidase—peroxidase method [J. Clin. Pathol. 22, 246 (1969)]. The reaction is linear for amylase activities up to 1400 Somogyi units/dl (2560 U/liter) and for maltose concentrations through 1500 mg/dl. No blank assay is required; consequently precision is improved and the automated system is simplified. Calibration with primary maltose standards increases accuracy and reliability. Common reducing substances in serum and urine do not interfere at their normal concentrations. There is a linear correlation between the results of this method and those of chromogenic and iodometric methods for normal and pathologic sera and urines. The chromogenic method yields significantly higher results and the iodometric method significantly lower results than this maltogenic method for elevated amylase activities. The normal range is 40-140 Somogyi units/dl (73-256 U/liter).

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