Evaluation of Kinetic Enzyme Parameters by Use of a Small Computer Interfaced "Fast Analyzer"—An Addition to Automated Clinical Enzymology

1973 ◽  
Vol 19 (8) ◽  
pp. 908-918 ◽  
Author(s):  
T O Tiffany ◽  
D D Chilcote ◽  
C A Burtis
Keyword(s):  
Alkaline Phosphatase ◽  
Kinetic Parameters ◽  
Small Computer ◽  
Programming Approach ◽  
Initial Reaction ◽  
Placental Alkaline Phosphatase ◽  
Intestinal Alkaline Phosphatase ◽  
On Line ◽  
Clinical Enzymology ◽  
Glucose 6 Phosphate Dehydrogenase

Abstract The evaluation of steady-state enzyme kinetic parameters such as Km, Vmax, and Ki is useful in clinical enzymology, particularly where they can be used for the detection of enzyme variants and to assess the type of isoenzymes and the extent of their abnormal activity. The multiple-cuvet parallel-analysis feature of the Fast Analyzer permits one to evaluate one or more enzymic parameters in a single rotor. Both on-line measurement of initial reaction velocities and rapid off-line reduction of statistical data can be obtained by using the small-computer interfaced Fast Analyzer. Estimates of error in determining these parameters are also obtained, for use in comparing results in future analyses. We discuss the programming approach for adapting both simple and more complex statistical nonlinear regression data-fitting routines to the Fast Analyzer’s computer. We illustrate how the analyzer may be used to determine simple kinetic parameters for placental alkaline phosphatase and glucose-6-phosphate dehydrogenase and for the more complex "stripping out" of Km’s and Vmax’s for a heterogenous mixture of placental and intestinal alkaline phosphatase.

Macromolecular alkaline phosphatase and an immunoglobulin G that inhibited alkaline phosphatase in a patient's serum.

1983 ◽  
Vol 29 (2) ◽  
pp. 375-378 ◽  
Author(s):  
H Nakagawa ◽  
K Umeki ◽  
K Yamanaka ◽  
N Kida ◽  
S Ohtaki
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Immunoglobulin G ◽  
Significant Loss ◽  
Placental Alkaline Phosphatase ◽  
Type Ii ◽  
Intestinal Alkaline Phosphatase ◽  
Adult Type ◽  
Heat Stable

Abstract Macromolecular alkaline phosphatase (EC 3.1.3.1) was found in the serum of a patient suffering from myasthenia gravis (adult type II) complicated with thymoma, and was shown by immunoelectrophoresis to be bound to immunoglobulins A and G (IgG). Placental alkaline phosphatase, complexed with either the patient's serum or IgG purified from the patient's serum, remained at the origin on electrophoresis, with significant loss of activity. Intestinal alkaline phosphatase, complexed with either the patient's serum or the patient's IgG, migrated to a position similar to that of the macromolecular alkaline phosphatase in the patient's serum on electrophoresis. About 50% of the placental alkaline phosphatase activity was inhibited with 0.1-0.2 g of the patient's IgG per liter, but 6.93 g of the IgG per liter was required for about 20% inhibition of the intestinal alkaline phosphatase activity. The complex of intestinal alkaline phosphatase with the patient's IgG was fairly heat stable. From these results, we concluded that the macromolecular alkaline phosphatase in the patient's serum consisted of intestinal alkaline phosphatase and IgG that was specific for placental alkaline phosphatase.

Placental alkaline phosphatase as a tumor marker for primary intracranial germinoma

1988 ◽  
Vol 68 (5) ◽  
pp. 710-720 ◽  
Author(s):  
Jun Shinoda ◽  
Hiromu Yamada ◽  
Noboru Sakai ◽  
Takashi Ando ◽  
Toshifumi Hirata ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Tumor Marker ◽  
Cross Reactivity ◽  
Cyst Fluid ◽  
Enzyme Linked Immunosorbent Assay ◽  
Placental Alkaline Phosphatase ◽  
Intestinal Alkaline Phosphatase ◽  
Content Type ◽  
Intracranial Germinoma ◽  
Very High

✓ A sensitive enzyme-linked immunosorbent assay (ELISA) was used in a retrospective study of placental alkaline phosphatase (PLAP) levels in serum, cerebrospinal fluid (CSF), and intratumoral cyst fluid in primary intracranial germinoma. The ELISA showed no cross-reactivity with intestinal alkaline phosphatase except in very high concentrations, after samples had been heat-treated. Three patients with germinoma were studied for serum PLAP levels and in all the levels were elevated (3.78, 0.52, and 2.11 IU/liter). Two of the germinoma patients were studied for PLAP levels in the CSF, and both had elevated levels (0.83 and 9.83 IU/liter). The intratumoral cyst fluid in one case of germinoma was tested for PLAP and the level was found to be very high (603 IU/liter). These PLAP levels decreased concomitantly with the reduction in tumor size during irradiation. Serum PLAP levels were measured in 40 control adult male individuals and in the CSF of 20 nonpregnant patients with subarachnoid hemorrhage. The upper normal limits were 0.20 and 0.11 IU/liter in the serum and the CSF, respectively. All PLAP levels measured in the serum of patients with various brain tumors were 0.18 IU/liter or less. This study strongly suggests that PLAP is a clinically useful tumor marker for primary intracranial germinoma.

scholarly journals Specific Immunoassays for Placental Alkaline Phosphatase As a Tumor Marker

2006 ◽  
Vol 2006 ◽  
pp. 1-8 ◽  
Author(s):  
Sérvio T. Stinghen ◽  
Juliana F. Moura ◽  
Patrícia Zancanella ◽  
Giovanna A. Rodrigues ◽  
Mara A. Pianovski ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Synthetic Peptide ◽  
Embryonal Carcinoma ◽  
Cross Reactivity ◽  
Placental Alkaline Phosphatase ◽  
Intestinal Alkaline Phosphatase ◽  
Alkaline Phosphatases ◽  
Cutoff Value ◽  
Heat Stable ◽  
Coefficients Of Variation

Human placental (hPLAP) and germ cell (PLAP-like) alkaline phosphatases are polymorphic and heat-stable enzymes. This study was designed to develop specific immunoassays for quantifying hPLAP and PLAP-like enzyme activity (EA) in sera of cancer patients, pregnant women, or smokers. Polyclonal sheep anti-hPLAP antibodies were purified by affinity chromatography with whole hPLAP protein (ICA-PLAP assay) or a synthetic peptide (aa 57–71) of hPLAP (ICA-PEP assay); the working range was0.1–11U/L and cutoff value was0.2U/L EA for nonsmokers. The intra- and interassay coefficients of variation were3.7%–6.5% (ICA-PLAP assay) and9.0%–9.9% (ICA-PEP assay). An insignificant cross-reactivity was noted for high levels of unheated intestinal alkaline phosphatase in ICA-PEP assay. A positive correlation between the regression of tumor size and EA was noted in a child with embryonal carcinoma. It can be concluded that ICA-PEP assay is more specific than ICA-PLAP, which is still useful to detect other PLAP/PLAP-like phenotypes.

Caco-2 cell transfection by rat intestinal alkaline phosphatase cDNA increases surfactant-like particles

1992 ◽  
Vol 263 (5) ◽  
pp. G756-G766 ◽  
Author(s):  
C. C. Tietze ◽  
M. J. Becich ◽  
M. Engle ◽  
W. F. Stenson ◽  
A. Mahmood ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Culture Medium ◽  
Lower Surface ◽  
Cell Transfection ◽  
Placental Alkaline Phosphatase ◽  
Intestinal Alkaline Phosphatase ◽  
Dipalmitoyl Phosphatidylcholine ◽  
Brush Borders ◽  
Human Placental Alkaline Phosphatase ◽  
Junctional Complexes

The rat enterocyte produces a particle with surfactant-like properties (including a whorled appearance, enrichment for dipalmitoyl phosphatidylcholine, and ability to lower surface tension) that also is enriched for intestinal alkaline phosphatase. Human Caco-2 cells grown on polycarbonate filters were utilized to study the secretion of these particles and exhibited whorls and strands of unilamellar membranes, particularly concentrated at the apical pole or near junctional complexes. Concentrated culture medium from these cells separated on continuous NaBr gradients revealed a fraction at density = 1.07 g/l enriched for phosphatidylcholine and intestinal alkaline phosphatase. This fraction contained membranous sheets containing alkaline phosphatase, detected by immunolocalization. Phosphatidylcholine comprised 54% of phospholipid in this fraction, compared with 20% in brush borders. When Caco-2 cells were transfected with cDNA encoding rat intestinal alkaline phosphatase, cellular phosphatase activity increased twofold, but activity in the medium increased 14-fold to > 200 (average 32)-fold. Ultrastructurally, compared with mock-transfected cells or cells transfected with human placental alkaline phosphatase, transfection with rat intestinal alkaline phosphatase cDNA led to intracellular and extracellular accumulation of surfactant-like particles. We conclude that surfactant-like particles are produced by Caco-2 cells, and their production can be enhanced by transfection with a cDNA encoding a protein known to be associated with such particles.

Image registration with a computer driven S.T.E.M.

1978 ◽  
Vol 36 (1) ◽  
pp. 98-99
Author(s):  
A.M.H. Schepman ◽  
J.A.P. van der Voort ◽  
J.E. Mellema
Keyword(s):  
Spot Size ◽  
Scanning Transmission Electron Microscope ◽  
Small Computer ◽  
Structural Studies ◽  
Area Of Interest ◽  
Transmission Electron ◽  
Scanning Transmission ◽  
Specimen Area ◽  
On Line ◽  
Minimum Distances

A Scanning Transmission Electron Microscope (STEM) was coupled to a small computer. The system (see Fig. 1) has been built using a Philips EM400, equipped with a scanning attachment and a DEC PDP11/34 computer with 34K memory. The gun (Fig. 2) consists of a continuously renewed tip of radius 0.2 to 0.4 μm of a tungsten wire heated just below its melting point by a focussed laser beam (1). On-line operation procedures were developped aiming at the reduction of the amount of radiation of the specimen area of interest, while selecting the various imaging parameters and upon registration of the information content. Whereas the theoretical limiting spot size is 0.75 nm (2), routine resolution checks showed minimum distances in the order 1.2 to 1.5 nm between corresponding intensity maxima in successive scans. This value is sufficient for structural studies of regular biological material to test the performance of STEM over high resolution CTEM.

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