Preparation of Entamoeba histolytica antigens without enzymatic inhibitors
The goal of this work is to report a novel assay that preserves antigenicity of extracts with high protease content without using enzymatic inhibitors. A great reduction of proteolytic activity in the insoluble chloroform/methanol and heated amoebic fraction (IC[ratio ]MC) was obtained by this metho...
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| Veröffentlicht in: | Parasitology 2005-08, Vol.131 (2), p.231-236 |
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| creator | FLORES, M. S. TAMEZ-TREVIÑO, E. CASTAÑEDA, F. TIJERINA-MENCHACA, R. GALAN-WONG, L. RANGEL, R. |
| description | The goal of this work is to report a novel assay that preserves antigenicity of extracts with high protease content without using enzymatic inhibitors. A great reduction of proteolytic activity in the insoluble chloroform/methanol and heated amoebic fraction (IC[ratio ]MC) was obtained by this method, even in the presence of sodium dodecyl sulphate and 2-mercaptoethanol. The substrates azo-casein and the hide powder azure were used to determine the reduction of proteolytic activity of IC[ratio ]MC compared with the activity of crude extract and crude extract with iodoacetamide. The IC[ratio ]MC SDS-PAGE pattern shows a higher quantity of bands than extract with the inhibitor iodoacetamide or than crude extract. In addition, anti-Entamoeba histolytica antibodies from amoebic liver abscess patients recognized a richer antigenic Western blot pattern in the IC[ratio ]MC fraction than in crude extract alone or with inhibitor. The described method has proved to be suitable to preserve amoebic antigens for its use in diagnostic tests and it can be used for immunological response studies against E. histolytica antigens. Furthermore we propose that this method to obtain the IC[ratio ]MC fraction can be applied for the study of other microorganisms or cells with high enzymatic content. |
| doi_str_mv | 10.1017/S0031182005007730 |
| format | Article |
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S. ; TAMEZ-TREVIÑO, E. ; CASTAÑEDA, F. ; TIJERINA-MENCHACA, R. ; GALAN-WONG, L. ; RANGEL, R.</creator><creatorcontrib>FLORES, M. S. ; TAMEZ-TREVIÑO, E. ; CASTAÑEDA, F. ; TIJERINA-MENCHACA, R. ; GALAN-WONG, L. ; RANGEL, R.</creatorcontrib><description>The goal of this work is to report a novel assay that preserves antigenicity of extracts with high protease content without using enzymatic inhibitors. A great reduction of proteolytic activity in the insoluble chloroform/methanol and heated amoebic fraction (IC[ratio ]MC) was obtained by this method, even in the presence of sodium dodecyl sulphate and 2-mercaptoethanol. The substrates azo-casein and the hide powder azure were used to determine the reduction of proteolytic activity of IC[ratio ]MC compared with the activity of crude extract and crude extract with iodoacetamide. The IC[ratio ]MC SDS-PAGE pattern shows a higher quantity of bands than extract with the inhibitor iodoacetamide or than crude extract. In addition, anti-Entamoeba histolytica antibodies from amoebic liver abscess patients recognized a richer antigenic Western blot pattern in the IC[ratio ]MC fraction than in crude extract alone or with inhibitor. The described method has proved to be suitable to preserve amoebic antigens for its use in diagnostic tests and it can be used for immunological response studies against E. histolytica antigens. Furthermore we propose that this method to obtain the IC[ratio ]MC fraction can be applied for the study of other microorganisms or cells with high enzymatic content.</description><identifier>ISSN: 0031-1820</identifier><identifier>EISSN: 1469-8161</identifier><identifier>DOI: 10.1017/S0031182005007730</identifier><identifier>PMID: 16145939</identifier><identifier>CODEN: PARAAE</identifier><language>eng</language><publisher>Cambridge, UK: Cambridge University Press</publisher><subject>Animals ; Antibodies, Protozoan - immunology ; antigen preservation ; Antigens, Protozoan - isolation & purification ; Antigens, Protozoan - metabolism ; Biological and medical sciences ; cellular enzymes ; Chloroform ; Diagnostic tests ; Entamoeba histolytica ; Entamoeba histolytica - enzymology ; Entamoeba histolytica - immunology ; enzymatic inhibitors ; Enzymes ; Fundamental and applied biological sciences. Psychology ; General aspects ; General aspects and techniques. Study of several systematic groups. Models ; Humans ; Invertebrates ; Liver Abscess, Amebic - immunology ; Methods ; Microorganisms ; Molecular weight ; Patients ; Peptide Hydrolases - metabolism ; proteolytic activity</subject><ispartof>Parasitology, 2005-08, Vol.131 (2), p.231-236</ispartof><rights>2005 Cambridge University Press</rights><rights>2005 INIST-CNRS</rights><rights>Copyright Cambridge University Press Aug 2005</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c438t-bb220c3bba3e4fd198ac42b3e1b7865e0cca79ee88c747ba85609feca86fcccb3</citedby><cites>FETCH-LOGICAL-c438t-bb220c3bba3e4fd198ac42b3e1b7865e0cca79ee88c747ba85609feca86fcccb3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.cambridge.org/core/product/identifier/S0031182005007730/type/journal_article$$EHTML$$P50$$Gcambridge$$H</linktohtml><link.rule.ids>164,314,780,784,27924,27925,55628</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=17024511$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/16145939$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>FLORES, M. S.</creatorcontrib><creatorcontrib>TAMEZ-TREVIÑO, E.</creatorcontrib><creatorcontrib>CASTAÑEDA, F.</creatorcontrib><creatorcontrib>TIJERINA-MENCHACA, R.</creatorcontrib><creatorcontrib>GALAN-WONG, L.</creatorcontrib><creatorcontrib>RANGEL, R.</creatorcontrib><title>Preparation of Entamoeba histolytica antigens without enzymatic inhibitors</title><title>Parasitology</title><addtitle>Parasitology</addtitle><description>The goal of this work is to report a novel assay that preserves antigenicity of extracts with high protease content without using enzymatic inhibitors. A great reduction of proteolytic activity in the insoluble chloroform/methanol and heated amoebic fraction (IC[ratio ]MC) was obtained by this method, even in the presence of sodium dodecyl sulphate and 2-mercaptoethanol. The substrates azo-casein and the hide powder azure were used to determine the reduction of proteolytic activity of IC[ratio ]MC compared with the activity of crude extract and crude extract with iodoacetamide. The IC[ratio ]MC SDS-PAGE pattern shows a higher quantity of bands than extract with the inhibitor iodoacetamide or than crude extract. In addition, anti-Entamoeba histolytica antibodies from amoebic liver abscess patients recognized a richer antigenic Western blot pattern in the IC[ratio ]MC fraction than in crude extract alone or with inhibitor. The described method has proved to be suitable to preserve amoebic antigens for its use in diagnostic tests and it can be used for immunological response studies against E. histolytica antigens. Furthermore we propose that this method to obtain the IC[ratio ]MC fraction can be applied for the study of other microorganisms or cells with high enzymatic content.</description><subject>Animals</subject><subject>Antibodies, Protozoan - immunology</subject><subject>antigen preservation</subject><subject>Antigens, Protozoan - isolation & purification</subject><subject>Antigens, Protozoan - metabolism</subject><subject>Biological and medical sciences</subject><subject>cellular enzymes</subject><subject>Chloroform</subject><subject>Diagnostic tests</subject><subject>Entamoeba histolytica</subject><subject>Entamoeba histolytica - enzymology</subject><subject>Entamoeba histolytica - immunology</subject><subject>enzymatic inhibitors</subject><subject>Enzymes</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>General aspects</subject><subject>General aspects and techniques. Study of several systematic groups. Models</subject><subject>Humans</subject><subject>Invertebrates</subject><subject>Liver Abscess, Amebic - immunology</subject><subject>Methods</subject><subject>Microorganisms</subject><subject>Molecular weight</subject><subject>Patients</subject><subject>Peptide Hydrolases - metabolism</subject><subject>proteolytic activity</subject><issn>0031-1820</issn><issn>1469-8161</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><recordid>eNp1kMFu1DAURS1ERacDH8AGRUh0l2LHie0soSoFVJWBlg0b69njdFwSe7AdwfD1OJqoI4FYeXHPfbo-CD0n-Ixgwl_fYEwJERXGDcacU_wILUjN2lIQRh6jxRSXU36MTmK8xxgzyqon6DjHddPSdoE-roLZQoBkvSt8V1y4BIM3CoqNjcn3u2Q1FOCSvTMuFj9t2vgxFcb93g25pAvrNlbZ5EN8io466KN5Nr9L9PXdxe35-_Lq0-WH8zdXpa6pSKVSVYU1VQqoqbs1aQXoulLUEMUFawzWGnhrjBCa11yBaBhuO6NBsE5rregSne7vboP_MZqY5GCjNn0PzvgxSjY1CGUZfPkXeO_H4PI2WWVNpJ0cLBHZQzr4GIPp5DbYAcJOEiwny_Ify7nzYj48qsGsD41ZawZezQBEDX0XwGkbDxzHVd0Qkrlyz2XX5tdDDuG7ZJzyRrLLz5J8W11_ub15K1eZp_NYGFSw6ztz-NL_5_4BnJ6j-Q</recordid><startdate>20050801</startdate><enddate>20050801</enddate><creator>FLORES, M. 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Psychology</topic><topic>General aspects</topic><topic>General aspects and techniques. Study of several systematic groups. Models</topic><topic>Humans</topic><topic>Invertebrates</topic><topic>Liver Abscess, Amebic - immunology</topic><topic>Methods</topic><topic>Microorganisms</topic><topic>Molecular weight</topic><topic>Patients</topic><topic>Peptide Hydrolases - metabolism</topic><topic>proteolytic activity</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>FLORES, M. 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S.</au><au>TAMEZ-TREVIÑO, E.</au><au>CASTAÑEDA, F.</au><au>TIJERINA-MENCHACA, R.</au><au>GALAN-WONG, L.</au><au>RANGEL, R.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Preparation of Entamoeba histolytica antigens without enzymatic inhibitors</atitle><jtitle>Parasitology</jtitle><addtitle>Parasitology</addtitle><date>2005-08-01</date><risdate>2005</risdate><volume>131</volume><issue>2</issue><spage>231</spage><epage>236</epage><pages>231-236</pages><issn>0031-1820</issn><eissn>1469-8161</eissn><coden>PARAAE</coden><abstract>The goal of this work is to report a novel assay that preserves antigenicity of extracts with high protease content without using enzymatic inhibitors. A great reduction of proteolytic activity in the insoluble chloroform/methanol and heated amoebic fraction (IC[ratio ]MC) was obtained by this method, even in the presence of sodium dodecyl sulphate and 2-mercaptoethanol. The substrates azo-casein and the hide powder azure were used to determine the reduction of proteolytic activity of IC[ratio ]MC compared with the activity of crude extract and crude extract with iodoacetamide. The IC[ratio ]MC SDS-PAGE pattern shows a higher quantity of bands than extract with the inhibitor iodoacetamide or than crude extract. In addition, anti-Entamoeba histolytica antibodies from amoebic liver abscess patients recognized a richer antigenic Western blot pattern in the IC[ratio ]MC fraction than in crude extract alone or with inhibitor. The described method has proved to be suitable to preserve amoebic antigens for its use in diagnostic tests and it can be used for immunological response studies against E. histolytica antigens. Furthermore we propose that this method to obtain the IC[ratio ]MC fraction can be applied for the study of other microorganisms or cells with high enzymatic content.</abstract><cop>Cambridge, UK</cop><pub>Cambridge University Press</pub><pmid>16145939</pmid><doi>10.1017/S0031182005007730</doi><tpages>6</tpages></addata></record> |
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| ispartof | Parasitology, 2005-08, Vol.131 (2), p.231-236 |
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| subjects | Animals Antibodies, Protozoan - immunology antigen preservation Antigens, Protozoan - isolation & purification Antigens, Protozoan - metabolism Biological and medical sciences cellular enzymes Chloroform Diagnostic tests Entamoeba histolytica Entamoeba histolytica - enzymology Entamoeba histolytica - immunology enzymatic inhibitors Enzymes Fundamental and applied biological sciences. Psychology General aspects General aspects and techniques. Study of several systematic groups. Models Humans Invertebrates Liver Abscess, Amebic - immunology Methods Microorganisms Molecular weight Patients Peptide Hydrolases - metabolism proteolytic activity |
| title | Preparation of Entamoeba histolytica antigens without enzymatic inhibitors |
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