Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.1 (alkaline phosphatase)
 47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Alkaline phosphatase has been localized ultracytochemically in PMN of man with normal and elevated levels of this enzyme. Contrary to guinea-pig PMN, no activity appears to be present in the specific granules. Instead, the plasma membrane and the membrane of the endocytic vacuoles show a strong staining. However, the demonstration of this activity depends on the preparatory procedure employed for PMN isolation. The use of dextran and Ficoll-Hypaque in the isolation procedure induces a marked increase in alkaline phosphatase staining of the PMN plasma membrane. Strongly increased activity at this site has been found in PMN from cancer patients. In most of them, additional staining has been observed in atypical vesicles and sometimes in the Golgi apparatus. These findings are discussed in the light of some previously reported controversial biochemical and cytochemical data on the distribution of alkaline phosphatase in human PMN.
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PMID:Alkaline phosphatase activity in human polymorphonuclear leukocytes. 62 Nov 60

Normal human granulocytes obtained by Ficoll-Hypaque sedimentation were subjected to mild hypotonic shock and disruption by shear. The homogenate was fractionated by differential centrifugation and equilibrium ultracentrifugation to yield a plasma membrane preparation constituting 1% of the total cellular protein and enriched fifteen- and six-fold in alkaline phosphatase and Mg2+-adenosine triphosphatase activities, respectively. Granulocytes obtained from patients with chronic myeloid leukemia (CML) were identically processed. The protein constituents of both the normal and CML granulocyte plasma membranes were resolved by two-dimensional polyacrylamide gel electrophoresis. Comparison of the stained gels revealed CML-associated quantitative changes in four out of the fifteen protein spots examined. Thus, this analysis has permitted identification of those protein moieties that deserve attention for further isolation and purification.
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PMID:Plasma membranes from normal and chronic myeloid leukemic granulocytes: isolation and two-dimensional polyacrylamide gel electrophoretic analysis. 385 66

Human blood neutrophilic leukocytes were separated and purified by modifications of the Hypaque/Ficoll and dextran separation methods, resulting in a suspension which was greater than 96% neutrophils. Neutrophils were prepared in 0.34 M sucrose containing heparin and were clarified of nongranular debris by sequential passage through polycarbonate filters of pore size 5 mu and 2 mu. Isopycnic sucrose gradients of such filtrates revealed three major bands. The gradient separated fractions were studied by electron microscopy including peroxidase cytochemistry and by enzyme assay for myeloperoxidase (MPO), beta-glucuronidase, muramidase alkaline phosphatase and acid phosphatase utilizing both p-nitrophenylphosphate (pnp) and beta-glycerophosphate as substrates. Peroxidase-positive granules were observed at both density 1.22 (band A) and density 1.20 (band B). Three peroxidase-negative granules were identified: the round or oval peroxidase-negative granule of density 1.22 (band A) and two smaller granules, distinguishable by size and shape at density 1.18 (band C). Band C granules contain crystalloid inclusions. Peaks of muramidase activity coincided with bands A and C, suggesting the presence of muramidase in the peroxidase-negative granules of density 1.22 and in one or both of the peroxidase-negative granules at density 1.18. beta-Glucuronidase was distributed like MPO, with a major peak in band B and a minor peak in band A. Acid beta-glycerophosphatase was largely in band A. Acid pnp phosphatase was nonspecifically associated with soluble nongranular protein which always remained at the origin of sucrose gradients. Alkaline phosphatase was not granule associated and sedimented alone to density 1.145, which is highly suggestive of a cytoplasmic membrane localization for this enzyme.
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PMID:Separation and characterization of human neutrophil granules. 444 23

Using electron microscope cytochemistry and cells separated on Ficoll-Hypaque, Mg2+-dependent ATPase, ADPase and 5'-nucleotidase were predominantly localized as ectoenzymes on normal human granulocytes. Large deposits of ATPase final reaction product and more finely granular deposits of 5'-nucleotidase final reaction product were firmly attached to the outer surface of cell plasma membranes. The final reaction product from ecto-ADPase was, however, only loosely associated with the plasma membrane. In addition, finer deposits of ADPase final reaction product were seen in specific granules and in background cytoplasm. No nucleotidase phosphatase activity was localized to the alkaline phosphatase-containing granules (phosphasomes) recently described by Rustin et al. In granulocytes from patients with chronic granulocytic leukaemia, ecto-ATPase had a patchy distribution on the plasma membranes. There was considerable heterogeneity between cells with regard to ADPase and 5'-nucleotidase localization. In some cells, ADPase was seen only at both site, while in some cells no activity was detected. 5'-Nucleotidase localization was normal in some cells but lacking from many. No correlation was found between enzyme heterogeneity and the degree of morphological cell maturity.
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PMID:Electron microscopic cytochemical localization of nucleoside phosphatases in normal and chronic granulocytic leukaemic human neutrophils. 611 13

Human lymphocytes were isolated from defibrinated blood by Ficoll-Hypaque centrifugation with erythrocyte hypotonic lysis. Homogenates of mixed lymphocytes were subjected to analytical subcellular fractionation by sucrose gradient centrifugation in a Beaufay automatic zonal rotor. The principal organelles were characterized by their marker enzymes: cytosol (lactate dehydrogenase), plasma membrane (5'-nucleotidase), endoplasmic reticulum (neutral alpha-glucosidase), mitochondria (malate dehydrogenase), lysosomes (N-acetyl-beta-glucosaminidase), peroxisomes (catalase). gamma-Glutamyl transferase was exclusively localized to the plasma membrane. Leucine amino-peptidase, especially when assayed in the presence of Co2+, was also partially localized to the plasma membrane. Experiments with diazotized sulphanilic acid, a non-permeant enzyme inhibitor, showed that these plasma membrane enzymes are present on the cell surface. No detectable alkaline phosphatase was found in the lymphocytes. Acid phosphatase and beta-glucuronidase were localized to lysosomes and there was some evidence for lysosomal heterogeneity. Leucine amino peptidase, optimal at pH 8.0, showed a partial localization to intracellular vesicles, possibly lysosomes, especially when assayed in the presence of EDTA. These studies provide a technique for determining the intracellular distribution of hitherto unassigned lymphocyte constituents and serve as a basis for investigating the cell pathology of lymphocytic disorders.
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PMID:Enzyme analysis and subcellular fractionation of human peripheral blood lymphocytes with special reference to the localization of putative plasma membrane enzymes. 614 55

Polymorphonuclear leukocytes were isolated from the peripheral blood of rabbits by Ficoll-Hypaque centrifugation followed by dextran sedimentation. The granulocytes were homogenized in isotonic sucrose and subjected to analytical subcellular fractionation by sucrose density gradient centrifugation. Leucine aminopeptidase, when assayed with L-leucine-7-amido-4-methyl-coumarin as substrate, showed a similar distribution to N-acetyl-beta-glucosaminidase and thus is localized to the tertiary granules. There was no leucine aminopeptidase associated with the plasma membrane of these cells. Further experiments with purified plasma membranes and inhibitor studies using diazotized sulphanilic acid further confirmed that leucine aminopeptidase had a purely intracellular localization. Vitamin B-12 binding protein showed a similar localization to alkaline phosphatase indicating that, as in human polymorphonuclear leukocytes, vitamin B-12 binding protein is located to the specific granules.
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PMID:Localization of leucine aminopeptidase and vitamin B-12 binding protein in rabbit peripheral blood polymorphonuclear leukocytes. 715 Jun 59

Cytochemistry was used to differentiate chicken heterophils and eosinophils in blood smears and cytospin preparations of isolated leukocytes. Cytochemical staining also was performed to evaluate any alteration of cell staining characteristics after heterophils had been isolated using discontinuous Ficoll-Hypaque gradients. Cytochemical staining reactions of heterophils and eosinophils were the same before and after discontinuous gradient cell separation. Characteristic positive reactions did not occur when chicken heterophils were stained with alkaline phosphatase, peroxidase, Sudan black B, acid phosphatase, alpha-naphthyl acetate esterase, naphthol AS-D chloroacetate esterase, periodic acid-Schiff, or Luna's eosinophil granule techniques. Chicken eosinophils stained positively when peroxidase, Sudan black B, acid phosphatase, and Luna methods were performed. In highly cellular cytospin preparations, peroxidase reactions readily distinguished the few contaminating eosinophils (< 1%) from the heterophils.
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PMID:Cytochemical staining characteristics of chicken heterophils and eosinophils. 1268 38