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A new one-step method for the cytochemical localization of ouabain-sensitive, potassium-dependent p-nitrophenylphosphatase activity. 1980

H Mayahara, and K Fujimoto, and T Ando, and K Ogawa

A new one-step method for the light and electron microscopic localization of the ouabain-sensitive, K-dependent p-nitrophenylphosphatase (K-NPPase) activity of the Na-K-ATPase complex is introduced. The incubation medium contains p-nitrophenylphosphate (NPP) as substrate, lead citrate as the capture reagent, and dimethylsulfoxide (DMSO) as an activator. It is usable at the optimal pH of the K-NPPase, which is about pH 9.0 in the presence of 25% of DMSO. The effects of fixation, lead concentration, and DMSO on the enzyme activity were studied using rat kidney as a test tissue. The fixation of tissues in a mixture of 2% paraformaldehyde and 0.5% glutaraldehyde for 60 min at 0 degrees--4 degrees C preserved 45% of the enzyme activity. In the absence of DMSO, lead citrate (4.0 mM) caused 82% inhibition of the enzyme activity in fixed tissue. However, the addition of DMSO (25%) caused about 3-fold activation of the remaining activity. Cytochemical demonstration of the ouabain-sensitive K-NPPase activity was successfully made by this method at both light and electron microscopic levels.

UI MeSH Term Description Entries
D007668 Kidney Body organ that filters blood for the secretion of URINE and that regulates ion concentrations. Kidneys
D007854 Lead A soft, grayish metal with poisonous salts; atomic number 82, atomic weight 207.2, symbol Pb.
D008297 Male Males
D008854 Microscopy, Electron Microscopy using an electron beam, instead of light, to visualize the sample, thereby allowing much greater magnification. The interactions of ELECTRONS with specimens are used to provide information about the fine structure of that specimen. In TRANSMISSION ELECTRON MICROSCOPY the reactions of the electrons that are transmitted through the specimen are imaged. In SCANNING ELECTRON MICROSCOPY an electron beam falls at a non-normal angle on the specimen and the image is derived from the reactions occurring above the plane of the specimen. Electron Microscopy
D009597 4-Nitrophenylphosphatase An enzyme that catalyzes the hydrolysis of nitrophenyl phosphates to nitrophenols. At acid pH it is probably ACID PHOSPHATASE (EC 3.1.3.2); at alkaline pH it is probably ALKALINE PHOSPHATASE (EC 3.1.3.1). EC 3.1.3.41. 4-Nitrophenyl Phosphatase,K+-NPPase,K-Dependent p-Nitrophenylphosphatase,K-p NPPase,Nitrophenyl Phosphatase,p-NPPase,p-Nitrophenylphosphatase,para-Nitrophenyl Phosphatase,para-Nitrophenylphosphatase,4 Nitrophenyl Phosphatase,4 Nitrophenylphosphatase,K Dependent p Nitrophenylphosphatase,K p NPPase,K+ NPPase,p NPPase,p Nitrophenylphosphatase,p-Nitrophenylphosphatase, K-Dependent,para Nitrophenyl Phosphatase,para Nitrophenylphosphatase
D010042 Ouabain A cardioactive glycoside consisting of rhamnose and ouabagenin, obtained from the seeds of Strophanthus gratus and other plants of the Apocynaceae; used like DIGITALIS. It is commonly used in cell biological studies as an inhibitor of the NA(+)-K(+)-EXCHANGING ATPASE. Acocantherin,G-Strophanthin,Acolongifloroside K,G Strophanthin
D010744 Phosphoric Monoester Hydrolases A group of hydrolases which catalyze the hydrolysis of monophosphoric esters with the production of one mole of orthophosphate. Phosphatase,Phosphatases,Phosphohydrolase,Phosphohydrolases,Phosphomonoesterase,Phosphomonoesterases,Phosphoric Monoester Hydrolase,Hydrolase, Phosphoric Monoester,Hydrolases, Phosphoric Monoester,Monoester Hydrolase, Phosphoric
D011188 Potassium An element in the alkali group of metals with an atomic symbol K, atomic number 19, and atomic weight 39.10. It is the chief cation in the intracellular fluid of muscle and other cells. Potassium ion is a strong electrolyte that plays a significant role in the regulation of fluid volume and maintenance of the WATER-ELECTROLYTE BALANCE.
D002951 Citrates Derivatives of CITRIC ACID.
D004121 Dimethyl Sulfoxide A highly polar organic liquid, that is used widely as a chemical solvent. Because of its ability to penetrate biological membranes, it is used as a vehicle for topical application of pharmaceuticals. It is also used to protect tissue during CRYOPRESERVATION. Dimethyl sulfoxide shows a range of pharmacological activity including analgesia and anti-inflammation. DMSO,Dimethyl Sulphoxide,Dimethylsulfoxide,Dimethylsulphinyl,Dimethylsulphoxide,Dimexide,Rheumabene,Rimso,Rimso 100,Rimso-50,Sclerosol,Sulfinylbis(methane),Rimso 50,Rimso50,Sulfoxide, Dimethyl,Sulphoxide, Dimethyl

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