Measurement of the internal pH of mast cell granules using microvolumetric fluorescence and isotopic techniques

The intragranular pH of isolated mast cell granules was measured. Because of the minute amounts of isolated granules available, two techniques were developed by modifying aminoacridine fluorescence and [ 14C]methylamine accumulation techniques to permit measurements with microliter sample volumes. G...

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Veröffentlicht in:Arch. Biochem. Biophys.; (United States) 1987-04, Vol.254 (1), p.222-233
Hauptverfasser: De Young, Mary Beth, Nemeth, Edward F., Scarpa, Antonio
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Nemeth, Edward F.
Scarpa, Antonio
description The intragranular pH of isolated mast cell granules was measured. Because of the minute amounts of isolated granules available, two techniques were developed by modifying aminoacridine fluorescence and [ 14C]methylamine accumulation techniques to permit measurements with microliter sample volumes. Granule purity was demonstrated by electron microscopy, ruthenium red exclusion, and biochemical (histamine, mast cell granule protease) analysis. The internal pH was determined to be 5.55 ± 0.06, indicating that the pH environment within mast cell granules is not significantly different from that of previously studied granule types (i.e., chromaffin, platelet, pancreatic islet, and pituitary granules). Collapse of the pH gradient by NH 4 + was demonstrated with both techniques. No evidence of Cl − OH − or specific cation/H + transport was found, and major chloride permeability could not be unequivocably demonstrated. Ca 2+ and Cl − at concentrations normally present extracellularly destabilized granules in the presence of NH 4 +, but this phenomenon does not necessarily indicate a role for these ions in the exocytotic release of granule contents from intact cells. The pH measurement techniques developed for investigating the properties of granules in mast cells may be useful for studying other granules that can be obtained only in limited quantities.
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School of Medicine, Cleveland, OH</creatorcontrib><description>The intragranular pH of isolated mast cell granules was measured. Because of the minute amounts of isolated granules available, two techniques were developed by modifying aminoacridine fluorescence and [ 14C]methylamine accumulation techniques to permit measurements with microliter sample volumes. Granule purity was demonstrated by electron microscopy, ruthenium red exclusion, and biochemical (histamine, mast cell granule protease) analysis. The internal pH was determined to be 5.55 ± 0.06, indicating that the pH environment within mast cell granules is not significantly different from that of previously studied granule types (i.e., chromaffin, platelet, pancreatic islet, and pituitary granules). Collapse of the pH gradient by NH 4 + was demonstrated with both techniques. No evidence of Cl − OH − or specific cation/H + transport was found, and major chloride permeability could not be unequivocably demonstrated. Ca 2+ and Cl − at concentrations normally present extracellularly destabilized granules in the presence of NH 4 +, but this phenomenon does not necessarily indicate a role for these ions in the exocytotic release of granule contents from intact cells. 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School of Medicine, Cleveland, OH</creatorcontrib><title>Measurement of the internal pH of mast cell granules using microvolumetric fluorescence and isotopic techniques</title><title>Arch. Biochem. Biophys.; (United States)</title><addtitle>Arch Biochem Biophys</addtitle><description>The intragranular pH of isolated mast cell granules was measured. Because of the minute amounts of isolated granules available, two techniques were developed by modifying aminoacridine fluorescence and [ 14C]methylamine accumulation techniques to permit measurements with microliter sample volumes. Granule purity was demonstrated by electron microscopy, ruthenium red exclusion, and biochemical (histamine, mast cell granule protease) analysis. The internal pH was determined to be 5.55 ± 0.06, indicating that the pH environment within mast cell granules is not significantly different from that of previously studied granule types (i.e., chromaffin, platelet, pancreatic islet, and pituitary granules). Collapse of the pH gradient by NH 4 + was demonstrated with both techniques. No evidence of Cl − OH − or specific cation/H + transport was found, and major chloride permeability could not be unequivocably demonstrated. Ca 2+ and Cl − at concentrations normally present extracellularly destabilized granules in the presence of NH 4 +, but this phenomenon does not necessarily indicate a role for these ions in the exocytotic release of granule contents from intact cells. The pH measurement techniques developed for investigating the properties of granules in mast cells may be useful for studying other granules that can be obtained only in limited quantities.</description><subject>550201 - Biochemistry- Tracer Techniques</subject><subject>AMINES</subject><subject>Aminoacridines</subject><subject>ANIMAL CELLS</subject><subject>ANIMALS</subject><subject>Applied sciences</subject><subject>BASIC BIOLOGICAL SCIENCES</subject><subject>CARBON 14 COMPOUNDS</subject><subject>Carbon Radioisotopes</subject><subject>CONNECTIVE TISSUE CELLS</subject><subject>Cytoplasmic Granules - metabolism</subject><subject>Exact sciences and technology</subject><subject>FLUORESCENCE</subject><subject>Hydrogen-Ion Concentration</subject><subject>Ions</subject><subject>ISOTOPE APPLICATIONS</subject><subject>LABELLED COMPOUNDS</subject><subject>LUMINESCENCE</subject><subject>Male</subject><subject>MAMMALS</subject><subject>MAST CELLS</subject><subject>Mast Cells - metabolism</subject><subject>MEASURING METHODS</subject><subject>METHYLAMINE</subject><subject>Methylamines - metabolism</subject><subject>ORGANIC COMPOUNDS</subject><subject>Other techniques and industries</subject><subject>PH VALUE</subject><subject>RATS</subject><subject>RESPONSE MODIFYING FACTORS</subject><subject>RODENTS</subject><subject>SOMATIC CELLS</subject><subject>Spectrometry, Fluorescence</subject><subject>TRACER TECHNIQUES</subject><subject>VERTEBRATES</subject><issn>0003-9861</issn><issn>1096-0384</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1987</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kUFrFTEUhYMo9dn6DxSCiOhi7M1kJplsClJaK7S46T7kZW7ayEzyTDIF_30nfY-3dBXI-e7lnHMJ-cDgOwMmzgGAN2oQ7OsgvykANTTsFdkwUKIBPnSvyeaIvCXvcv4DwFgn2hNywnupWqU2JN6hyUvCGUOh0dHyiNSHgimYie5u6tdscqEWp4k-JBOWCTNdsg8PdPY2xac4LTOW5C110xITZovBIjVhpD7HEnerUtA-Bv93wXxG3jgzZXx_eE_J_fXV_eVNc_v756_LH7eN7aQqzWg6AdL1wFvgzHHRcdY7M4reWOUApdwCguud5a3BsTMWhOlsK5QYt6Lnp-TTfm3MxetsfXVgYwhoixYdqFYMK_RlD-1SrN6Knn2uQU3AuGQtZc-4egG7PbjmzTmh07vkZ5P-aQa63kLXonUtWg9Sv9xCs3Xs42H_sp1xPA4dyl_1zwfdZGsmt7ZrfT5ishfQ9nzFLvYYrn09eUw1T-149KnGGaP_v49n5UKm2Q</recordid><startdate>19870401</startdate><enddate>19870401</enddate><creator>De Young, Mary Beth</creator><creator>Nemeth, Edward F.</creator><creator>Scarpa, Antonio</creator><general>Elsevier Inc</general><general>Elsevier</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>OTOTI</scope></search><sort><creationdate>19870401</creationdate><title>Measurement of the internal pH of mast cell granules using microvolumetric fluorescence and isotopic techniques</title><author>De Young, Mary Beth ; Nemeth, Edward F. ; Scarpa, Antonio</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c479t-da4607f5032031f364315fad65ac9f0e77b0e0f5fc32aed4ac06a4c2696db653</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1987</creationdate><topic>550201 - Biochemistry- Tracer Techniques</topic><topic>AMINES</topic><topic>Aminoacridines</topic><topic>ANIMAL CELLS</topic><topic>ANIMALS</topic><topic>Applied sciences</topic><topic>BASIC BIOLOGICAL SCIENCES</topic><topic>CARBON 14 COMPOUNDS</topic><topic>Carbon Radioisotopes</topic><topic>CONNECTIVE TISSUE CELLS</topic><topic>Cytoplasmic Granules - metabolism</topic><topic>Exact sciences and technology</topic><topic>FLUORESCENCE</topic><topic>Hydrogen-Ion Concentration</topic><topic>Ions</topic><topic>ISOTOPE APPLICATIONS</topic><topic>LABELLED COMPOUNDS</topic><topic>LUMINESCENCE</topic><topic>Male</topic><topic>MAMMALS</topic><topic>MAST CELLS</topic><topic>Mast Cells - metabolism</topic><topic>MEASURING METHODS</topic><topic>METHYLAMINE</topic><topic>Methylamines - metabolism</topic><topic>ORGANIC COMPOUNDS</topic><topic>Other techniques and industries</topic><topic>PH VALUE</topic><topic>RATS</topic><topic>RESPONSE MODIFYING FACTORS</topic><topic>RODENTS</topic><topic>SOMATIC CELLS</topic><topic>Spectrometry, Fluorescence</topic><topic>TRACER TECHNIQUES</topic><topic>VERTEBRATES</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>De Young, Mary Beth</creatorcontrib><creatorcontrib>Nemeth, Edward F.</creatorcontrib><creatorcontrib>Scarpa, Antonio</creatorcontrib><creatorcontrib>Case Western Reserve Univ. 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Biophys.; (United States)</jtitle><addtitle>Arch Biochem Biophys</addtitle><date>1987-04-01</date><risdate>1987</risdate><volume>254</volume><issue>1</issue><spage>222</spage><epage>233</epage><pages>222-233</pages><issn>0003-9861</issn><eissn>1096-0384</eissn><coden>ABBIA4</coden><abstract>The intragranular pH of isolated mast cell granules was measured. Because of the minute amounts of isolated granules available, two techniques were developed by modifying aminoacridine fluorescence and [ 14C]methylamine accumulation techniques to permit measurements with microliter sample volumes. Granule purity was demonstrated by electron microscopy, ruthenium red exclusion, and biochemical (histamine, mast cell granule protease) analysis. The internal pH was determined to be 5.55 ± 0.06, indicating that the pH environment within mast cell granules is not significantly different from that of previously studied granule types (i.e., chromaffin, platelet, pancreatic islet, and pituitary granules). Collapse of the pH gradient by NH 4 + was demonstrated with both techniques. No evidence of Cl − OH − or specific cation/H + transport was found, and major chloride permeability could not be unequivocably demonstrated. Ca 2+ and Cl − at concentrations normally present extracellularly destabilized granules in the presence of NH 4 +, but this phenomenon does not necessarily indicate a role for these ions in the exocytotic release of granule contents from intact cells. The pH measurement techniques developed for investigating the properties of granules in mast cells may be useful for studying other granules that can be obtained only in limited quantities.</abstract><cop>San Diego, CA</cop><pub>Elsevier Inc</pub><pmid>3579299</pmid><doi>10.1016/0003-9861(87)90098-1</doi><tpages>12</tpages></addata></record>
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subjects 550201 - Biochemistry- Tracer Techniques
AMINES
Aminoacridines
ANIMAL CELLS
ANIMALS
Applied sciences
BASIC BIOLOGICAL SCIENCES
CARBON 14 COMPOUNDS
Carbon Radioisotopes
CONNECTIVE TISSUE CELLS
Cytoplasmic Granules - metabolism
Exact sciences and technology
FLUORESCENCE
Hydrogen-Ion Concentration
Ions
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
LUMINESCENCE
Male
MAMMALS
MAST CELLS
Mast Cells - metabolism
MEASURING METHODS
METHYLAMINE
Methylamines - metabolism
ORGANIC COMPOUNDS
Other techniques and industries
PH VALUE
RATS
RESPONSE MODIFYING FACTORS
RODENTS
SOMATIC CELLS
Spectrometry, Fluorescence
TRACER TECHNIQUES
VERTEBRATES
title Measurement of the internal pH of mast cell granules using microvolumetric fluorescence and isotopic techniques
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