A sensitive, simple assay of mitochondrial ATP synthesis of cultured mammalian cells suitable for high-throughput analysis
► A sensitive, simple assay for mitochondrial ATP synthesis that represents mitochondrial functional integrity. ► It needs no detergent and no mitochondrial fractionation. ► Good for high-throughput analysis of mammalian cells cultured on a microplate. A new assay has been developed to measure mitoc...
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| Veröffentlicht in: | Biochemical and biophysical research communications 2010-10, Vol.401 (4), p.538-543 |
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| creator | Fujikawa, Makoto Yoshida, Masasuke |
| description | ► A sensitive, simple assay for mitochondrial ATP synthesis that represents mitochondrial functional integrity. ► It needs no detergent and no mitochondrial fractionation. ► Good for high-throughput analysis of mammalian cells cultured on a microplate.
A new assay has been developed to measure mitochondrial ATP synthesis of cultured mammalian cells. Cells in a microplate are exposed to streptolysin O to make plasma membranes permeable without damaging mitochondrial function and ATP synthesis is monitored by luciferase. Addition of inhibitors of F
oF
1-ATP synthase (F
oF
1), respiratory chain, TCA cycle and ATP/ADP translocator, as well as knockdown of β-subunit of F
oF
1, resulted in loss of ATP synthesis. Compared with the conventional procedures that need mitochondria fractionation and detergent, this assay is simple, sensitive and suitable for high-throughput analysis of genes and drugs that could affect mitochondrial functional integrity as represented by ATP synthesis activity. |
| doi_str_mv | 10.1016/j.bbrc.2010.09.089 |
| format | Article |
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A new assay has been developed to measure mitochondrial ATP synthesis of cultured mammalian cells. Cells in a microplate are exposed to streptolysin O to make plasma membranes permeable without damaging mitochondrial function and ATP synthesis is monitored by luciferase. Addition of inhibitors of F
oF
1-ATP synthase (F
oF
1), respiratory chain, TCA cycle and ATP/ADP translocator, as well as knockdown of β-subunit of F
oF
1, resulted in loss of ATP synthesis. Compared with the conventional procedures that need mitochondria fractionation and detergent, this assay is simple, sensitive and suitable for high-throughput analysis of genes and drugs that could affect mitochondrial functional integrity as represented by ATP synthesis activity.</description><identifier>ISSN: 0006-291X</identifier><identifier>EISSN: 1090-2104</identifier><identifier>DOI: 10.1016/j.bbrc.2010.09.089</identifier><identifier>PMID: 20875793</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Adenosine Triphosphate - analysis ; Adenosine Triphosphate - biosynthesis ; Adenosine Triphosphate - genetics ; Animals ; ATP synthesis ; Bacterial Proteins - pharmacology ; Cell Line ; Citric Acid Cycle - drug effects ; F oF 1-ATP synthase ; Gene Knockdown Techniques ; High-throughput analysis ; High-Throughput Screening Assays ; Humans ; Mitochondria ; Mitochondria - metabolism ; Permeability ; Proton-Translocating ATPases - antagonists & inhibitors ; Proton-Translocating ATPases - genetics ; Sensitivity and Specificity ; Streptolysin ; Streptolysins - pharmacology</subject><ispartof>Biochemical and biophysical research communications, 2010-10, Vol.401 (4), p.538-543</ispartof><rights>2010 Elsevier Inc.</rights><rights>Copyright © 2010 Elsevier Inc. All rights reserved.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c421t-f5e6cafec09646e40eaa2dd7d61363024615565fd0ec9ff80b0943f17c8d67b13</citedby><cites>FETCH-LOGICAL-c421t-f5e6cafec09646e40eaa2dd7d61363024615565fd0ec9ff80b0943f17c8d67b13</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0006291X10017997$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,776,780,3537,27901,27902,65306</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/20875793$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Fujikawa, Makoto</creatorcontrib><creatorcontrib>Yoshida, Masasuke</creatorcontrib><title>A sensitive, simple assay of mitochondrial ATP synthesis of cultured mammalian cells suitable for high-throughput analysis</title><title>Biochemical and biophysical research communications</title><addtitle>Biochem Biophys Res Commun</addtitle><description>► A sensitive, simple assay for mitochondrial ATP synthesis that represents mitochondrial functional integrity. ► It needs no detergent and no mitochondrial fractionation. ► Good for high-throughput analysis of mammalian cells cultured on a microplate.
A new assay has been developed to measure mitochondrial ATP synthesis of cultured mammalian cells. Cells in a microplate are exposed to streptolysin O to make plasma membranes permeable without damaging mitochondrial function and ATP synthesis is monitored by luciferase. Addition of inhibitors of F
oF
1-ATP synthase (F
oF
1), respiratory chain, TCA cycle and ATP/ADP translocator, as well as knockdown of β-subunit of F
oF
1, resulted in loss of ATP synthesis. Compared with the conventional procedures that need mitochondria fractionation and detergent, this assay is simple, sensitive and suitable for high-throughput analysis of genes and drugs that could affect mitochondrial functional integrity as represented by ATP synthesis activity.</description><subject>Adenosine Triphosphate - analysis</subject><subject>Adenosine Triphosphate - biosynthesis</subject><subject>Adenosine Triphosphate - genetics</subject><subject>Animals</subject><subject>ATP synthesis</subject><subject>Bacterial Proteins - pharmacology</subject><subject>Cell Line</subject><subject>Citric Acid Cycle - drug effects</subject><subject>F oF 1-ATP synthase</subject><subject>Gene Knockdown Techniques</subject><subject>High-throughput analysis</subject><subject>High-Throughput Screening Assays</subject><subject>Humans</subject><subject>Mitochondria</subject><subject>Mitochondria - metabolism</subject><subject>Permeability</subject><subject>Proton-Translocating ATPases - antagonists & inhibitors</subject><subject>Proton-Translocating ATPases - genetics</subject><subject>Sensitivity and Specificity</subject><subject>Streptolysin</subject><subject>Streptolysins - pharmacology</subject><issn>0006-291X</issn><issn>1090-2104</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2010</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kE2LFDEQhoMo7rj6BzxIbl7ssZLuTnfAy7D4BQt6WMFbSCeV7Qz9MabSC-Ovt5tZPXoqqHrroeph7LWAvQCh3h_3XZfcXsLaAL2HVj9hOwEaCimgesp2AKAKqcXPK_aC6AggRKX0c3YloW3qRpc79vvACSeKOT7gO05xPA3ILZE98znwMebZ9fPkU7QDP9x953Seco8UaRu7ZchLQs9HO452iHbiDoeBOC0x224lhTnxPt73Re7TvNz3pyVzO9nhvBJesmfBDoSvHus1-_Hp493Nl-L22-evN4fbwlVS5CLUqJwN6ECrSmEFaK30vvFKlKoEWSlR16oOHtDpEFroQFdlEI1rvWo6UV6ztxfuKc2_FqRsxkjbnXbCeSHTKAlS61atSXlJujQTJQzmlOJo09kIMJtyczSbcrMpN6DNqnxdevOIX7oR_b-Vv47XwIdLANcnHyImQy7i5NDHhC4bP8f_8f8AvEyU8w</recordid><startdate>20101029</startdate><enddate>20101029</enddate><creator>Fujikawa, Makoto</creator><creator>Yoshida, Masasuke</creator><general>Elsevier Inc</general><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></search><sort><creationdate>20101029</creationdate><title>A sensitive, simple assay of mitochondrial ATP synthesis of cultured mammalian cells suitable for high-throughput analysis</title><author>Fujikawa, Makoto ; Yoshida, Masasuke</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c421t-f5e6cafec09646e40eaa2dd7d61363024615565fd0ec9ff80b0943f17c8d67b13</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2010</creationdate><topic>Adenosine Triphosphate - analysis</topic><topic>Adenosine Triphosphate - biosynthesis</topic><topic>Adenosine Triphosphate - genetics</topic><topic>Animals</topic><topic>ATP synthesis</topic><topic>Bacterial Proteins - pharmacology</topic><topic>Cell Line</topic><topic>Citric Acid Cycle - drug effects</topic><topic>F oF 1-ATP synthase</topic><topic>Gene Knockdown Techniques</topic><topic>High-throughput analysis</topic><topic>High-Throughput Screening Assays</topic><topic>Humans</topic><topic>Mitochondria</topic><topic>Mitochondria - metabolism</topic><topic>Permeability</topic><topic>Proton-Translocating ATPases - antagonists & inhibitors</topic><topic>Proton-Translocating ATPases - genetics</topic><topic>Sensitivity and Specificity</topic><topic>Streptolysin</topic><topic>Streptolysins - pharmacology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Fujikawa, Makoto</creatorcontrib><creatorcontrib>Yoshida, Masasuke</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Biochemical and biophysical research communications</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Fujikawa, Makoto</au><au>Yoshida, Masasuke</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A sensitive, simple assay of mitochondrial ATP synthesis of cultured mammalian cells suitable for high-throughput analysis</atitle><jtitle>Biochemical and biophysical research communications</jtitle><addtitle>Biochem Biophys Res Commun</addtitle><date>2010-10-29</date><risdate>2010</risdate><volume>401</volume><issue>4</issue><spage>538</spage><epage>543</epage><pages>538-543</pages><issn>0006-291X</issn><eissn>1090-2104</eissn><abstract>► A sensitive, simple assay for mitochondrial ATP synthesis that represents mitochondrial functional integrity. ► It needs no detergent and no mitochondrial fractionation. ► Good for high-throughput analysis of mammalian cells cultured on a microplate.
A new assay has been developed to measure mitochondrial ATP synthesis of cultured mammalian cells. Cells in a microplate are exposed to streptolysin O to make plasma membranes permeable without damaging mitochondrial function and ATP synthesis is monitored by luciferase. Addition of inhibitors of F
oF
1-ATP synthase (F
oF
1), respiratory chain, TCA cycle and ATP/ADP translocator, as well as knockdown of β-subunit of F
oF
1, resulted in loss of ATP synthesis. Compared with the conventional procedures that need mitochondria fractionation and detergent, this assay is simple, sensitive and suitable for high-throughput analysis of genes and drugs that could affect mitochondrial functional integrity as represented by ATP synthesis activity.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>20875793</pmid><doi>10.1016/j.bbrc.2010.09.089</doi><tpages>6</tpages></addata></record> |
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| ispartof | Biochemical and biophysical research communications, 2010-10, Vol.401 (4), p.538-543 |
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| language | eng |
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| source | MEDLINE; Elsevier ScienceDirect Journals |
| subjects | Adenosine Triphosphate - analysis Adenosine Triphosphate - biosynthesis Adenosine Triphosphate - genetics Animals ATP synthesis Bacterial Proteins - pharmacology Cell Line Citric Acid Cycle - drug effects F oF 1-ATP synthase Gene Knockdown Techniques High-throughput analysis High-Throughput Screening Assays Humans Mitochondria Mitochondria - metabolism Permeability Proton-Translocating ATPases - antagonists & inhibitors Proton-Translocating ATPases - genetics Sensitivity and Specificity Streptolysin Streptolysins - pharmacology |
| title | A sensitive, simple assay of mitochondrial ATP synthesis of cultured mammalian cells suitable for high-throughput analysis |
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