Serum Alkaline Phosphatase: Total Activity and Isoenzyme Determinations Made by Use of the Centrifugal Fast Analyzer

1972 ◽  
Vol 18 (12) ◽  
pp. 1468-1474 ◽  
Author(s):  
Bernard E Statland ◽  
H Harold Nishi ◽  
D S Young
Keyword(s):  
Alkaline Phosphatase ◽  
Clinical Laboratory ◽  
Serum Alkaline Phosphatase ◽  
Kinetic Method ◽  
Total Activity ◽  
Alkaline Phosphatase Isoenzymes ◽  
Chemical Inhibition ◽  
Routine Clinical Laboratory ◽  
Adult Volunteers ◽  
Total Alkaline

Abstract We present a kinetic method in which the centrifugal analyzer is used to determine total alkaline phosphatase activity and to measure the isoenzymes in bone, liver, and the L-phenylalanine-sensitive fraction (intestine, tumor, and placenta). Measurements were made at 30°C in diethanolamine buffer, with p-nitrophenylphosphate as substrate. The alkaline phosphatase isoenzymes were separated and measured by selective chemical inhibition with 10 mmol/liter L-phenylalanine and 3.3 mol urea per liter. Analyses were performed on sera of a group of healthy pediatric and young adult volunteers and, in addition, on sera from patients with clinically documented osteoblastic disorders and hepatobiliary diseases. The precision and accuracy of the centrifugal analyzer were evaluated, as was the suitability of the instrument for routine clinical laboratory use.

Quantitative alkaline phosphatase isoenzyme determination by electrophoresis on cellulose acetate membranes.

1977 ◽  
Vol 23 (1) ◽  
pp. 28-34 ◽  
Author(s):  
W H Siede ◽  
U B Seiffert
Keyword(s):  
Alkaline Phosphatase ◽  
Cellulose Acetate ◽  
Clinical Laboratory ◽  
Kinetic Assay ◽  
Specific Staining ◽  
Cellulose Acetate Membranes ◽  
Alkaline Phosphatase Isoenzymes ◽  
Elution Method ◽  
Routine Clinical Laboratory ◽  
Alkaline Phosphatase Isoenzyme

Abstract We present a new method for quantitative determination of alkaline phosphatase isoenzymes. This method consists of electrophoretic separation on cellulose acetate membranes, special fixation technique to avoid elution and diffusion of enzyme protein during incubation, specific staining, and quantitative evaluation by densitometric measurement. We highly recommend the precedure for routine clinical laboratory use. In all normal individuals we observe two isoenzymes of hepatic origin and one isoenzyme each of osseous, intestinal, and biliary origin. Quantitative normal values are presented. Precision of the method is calculated, the CV being less than 10%. The exactness of densitometric quantification is proved by comparison with kinetic assay of alkaline phosphatase isoenzymes by use of an elution method. Clinical implications of alkaline phosphatase isoenzymograms are reported and discussed in detail.

Hypophosphatasia (adult form): quantitation of serum alkaline phosphatase isoenzyme activity in a large kindred.

1980 ◽  
Vol 26 (7) ◽  
pp. 840-845 ◽  
Author(s):  
J L Millán ◽  
M P Whyte ◽  
L V Avioli ◽  
W H Fishman
Keyword(s):  
Alkaline Phosphatase ◽  
Serum Alkaline Phosphatase ◽  
Hepatic Metabolism ◽  
Total Activity ◽  
Heat Inactivation ◽  
Mean Values ◽  
Adult Form ◽  
Cellulose Acetate Membranes ◽  
Alkaline Phosphatase Isoenzymes ◽  
Alkaline Phosphatase Isoenzyme

Abstract We used heat inactivation, L-phenylalanine inhibition, and electrophoresis on polyacrylamide gel and cellulose acetate membranes--with and without use of specific antisera against the liver-bone, intestinal, and placental isoenzymes--to distinguish and quantitate the different alkaline phosphatase isoenzymes in sera from 23 adult members of a kindred affected by the adult form of hypophosphatasia. Nine subjects had values for total activity more than two standard deviations below the mean values for age- and sex-matched normal persons. Bone isoenzyme was diminished in all nine, whereas liver isoenzyme was subnormal in only four. Phosphoethanolamine and phosphoserine in the urine of eight hypophosphatasemic individuals correlated inversely with both total and liver alkaline phosphatase activity in their serum, but not with the activity of the bone isoenzyme. Total activity in the serum of adult kindred members correlated best with the circulating liver isoenzyme activity. The findings suggest that altered hepatic metabolism is responsible for the increased urinary excretion of phosphoethanolamine, and perhaps phosphoserine, in hypophosphatasia.

Electrophoretic Separation of Alkaline Phosphatase Isoenzymes: A Clinical Evaluation

1972 ◽  
Vol 17 (5) ◽  
pp. 172-175 ◽  
Author(s):  
R. R. G. Warwick ◽  
D. J. C. Shearman ◽  
I. W. Percy-Robb ◽  
A. F. Smith
Keyword(s):  
Alkaline Phosphatase ◽  
Gel Electrophoresis ◽  
Serum Alkaline Phosphatase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Total Serum ◽  
Electrophoretic Separation ◽  
Nucleotidase Activity ◽  
Total Alkaline Phosphatase ◽  
Alkaline Phosphatase Isoenzymes ◽  
Total Alkaline

In 40 patients with a raised total serum alkaline phosphatase, polyacrylamide gel electrophoresis was used to separate the liver, bone and intestinal isoenzymes of alkaline phosphatase. The method confirmed the source of the raised alkaline phosphatase in patients with established bone or liver disease. The technique was found to be more reliable than the estimation of 5'-nucleotidase activity in the detection of a raised alkaline phosphatase of hepatic origin and was also useful when the raised total alkaline phosphatase came from two sources.

Phenolphthalein Monophosphate as a Substrate for Serum Alkaline Phosphatase

1966 ◽  
Vol 12 (10) ◽  
pp. 701-708 ◽  
Author(s):  
J H Wilkinson ◽  
Ann V Vodden
Keyword(s):  
Alkaline Phosphatase ◽  
Serum Concentration ◽  
Inorganic Phosphate ◽  
Clinical Laboratory ◽  
Serum Alkaline Phosphatase ◽  
Routine Procedure ◽  
Laboratory Conditions ◽  
Straight Line ◽  
Routine Clinical Laboratory

Abstract The procedure for serum alkaline phosphatase determination employing phenolphthalein monophosphate as substrate was evaluated under routine clinical laboratory conditions. A straight-line response to increasing serum concentration was observed, and the reaction was shown to be Mg++ dependent. The method proved to be extremely simple, and the results obtained correlated well with those given by a standard routine procedure involving determination of the liberated inorganic phosphate.

Hypophosphatasia (adult form): quantitation of serum alkaline phosphatase isoenzyme activity in a large kindred.

1980 ◽  
Vol 26 (7) ◽  
pp. 840-845
Author(s):  
J L Millán ◽  
M P Whyte ◽  
L V Avioli ◽  
W H Fishman
Keyword(s):  
Alkaline Phosphatase ◽  
Serum Alkaline Phosphatase ◽  
Hepatic Metabolism ◽  
Total Activity ◽  
Heat Inactivation ◽  
Mean Values ◽  
Adult Form ◽  
Cellulose Acetate Membranes ◽  
Alkaline Phosphatase Isoenzymes ◽  
Alkaline Phosphatase Isoenzyme

Abstract We used heat inactivation, L-phenylalanine inhibition, and electrophoresis on polyacrylamide gel and cellulose acetate membranes--with and without use of specific antisera against the liver-bone, intestinal, and placental isoenzymes--to distinguish and quantitate the different alkaline phosphatase isoenzymes in sera from 23 adult members of a kindred affected by the adult form of hypophosphatasia. Nine subjects had values for total activity more than two standard deviations below the mean values for age- and sex-matched normal persons. Bone isoenzyme was diminished in all nine, whereas liver isoenzyme was subnormal in only four. Phosphoethanolamine and phosphoserine in the urine of eight hypophosphatasemic individuals correlated inversely with both total and liver alkaline phosphatase activity in their serum, but not with the activity of the bone isoenzyme. Total activity in the serum of adult kindred members correlated best with the circulating liver isoenzyme activity. The findings suggest that altered hepatic metabolism is responsible for the increased urinary excretion of phosphoethanolamine, and perhaps phosphoserine, in hypophosphatasia.

scholarly journals Reaction Rate Retardation as a Method for Serum Alkaline Phosphatase Isoenzyme Measurement

1980 ◽  
Vol 17 (4) ◽  
pp. 192-198 ◽  
Author(s):  
Pamela B Brown ◽  
K O Lewis
Keyword(s):  
Alkaline Phosphatase ◽  
Reaction Rate ◽  
Serum Alkaline Phosphatase ◽  
Enzyme Reaction ◽  
Intestinal Alkaline Phosphatase ◽  
Wide Range ◽  
Alkaline Phosphatase Isoenzymes ◽  
Sensitivity And Selectivity ◽  
Rate Retardation ◽  
Alkaline Phosphatase Isoenzyme

A method for serum alkaline phosphatase isoenzymes using an enzyme reaction rate analyser is described. The complete urea-induced degradation of enzyme activity is monitored, from which individual isoenzyme activities are obtained by calculating the constituent exponential components of the degradation curve. Activities have been measured with adequate sensitivity and selectivity for up to four isoenzyme components in normal and in pathological sera. The identity of each isoenzyme present is assigned from its characteristic degradation half-life, and by this method bone and liver alkaline phosphatase are clearly distinguished and quantitated, and a composite value for placental-intestinal alkaline phosphatase activity is obtained. The approach promises to be applicable to a wide range of isoenzymes, and in analogy with ‘reaction rate’ the term ‘reaction rate retardation’ is suggested for the procedure.

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