Subpopulations of rat lung fibroblasts with different amounts of type I and type III collagen mRNAs
A fluorescence-based method using the cell sorter has been devised to separate rat lung fibroblasts into subpopulations. Type I or type III collagen antiserum was used as the primary antibody to react with parent rat lung fibroblasts. This was followed by a fluorescein-conjugated secondary antibody....
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| Veröffentlicht in: | The Journal of biological chemistry 1990-04, Vol.265 (11), p.6286-6290 |
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| container_title | The Journal of biological chemistry |
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| creator | BREEN, E FALCO, V. M ABSHER, M CUTRONEO, K. R |
| description | A fluorescence-based method using the cell sorter has been devised to separate rat lung fibroblasts into subpopulations. Type
I or type III collagen antiserum was used as the primary antibody to react with parent rat lung fibroblasts. This was followed
by a fluorescein-conjugated secondary antibody. Specificity of the primary collagen antibody was determined using a monoclonal
beta-actin antibody and purified IgG as the primary antibodies. The fluorescent shift of parent rat lung fibroblasts was optimized
for the amount of primary collagen antibody and secondary fluorescein-conjugated antibody. An increase in slot blot intensity
was observed for pro-alpha 1(I), pro-alpha 2(I), and pro-alpha 1(III) mRNAs with increasing amounts of cellular RNA. When
precipitating with type I collagen antibodies, the total cellular steady-state levels of type I procollagen mRNAs were increased
in the high intensity cells as compared with the low intensity cells. Alternately, when the type III collagen antibodies were
used to precipitate the rat lung fibroblasts, the low intensity cells had increased type I procollagen mRNAs while the high
intensity cells had increased type III procollagen mRNA. The subpopulations of rat lung fibroblasts after isolation using
the fluorescent cell sorter were readily propagated for at least four passages. |
| doi_str_mv | 10.1016/S0021-9258(19)39323-8 |
| format | Article |
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I or type III collagen antiserum was used as the primary antibody to react with parent rat lung fibroblasts. This was followed
by a fluorescein-conjugated secondary antibody. Specificity of the primary collagen antibody was determined using a monoclonal
beta-actin antibody and purified IgG as the primary antibodies. The fluorescent shift of parent rat lung fibroblasts was optimized
for the amount of primary collagen antibody and secondary fluorescein-conjugated antibody. An increase in slot blot intensity
was observed for pro-alpha 1(I), pro-alpha 2(I), and pro-alpha 1(III) mRNAs with increasing amounts of cellular RNA. When
precipitating with type I collagen antibodies, the total cellular steady-state levels of type I procollagen mRNAs were increased
in the high intensity cells as compared with the low intensity cells. Alternately, when the type III collagen antibodies were
used to precipitate the rat lung fibroblasts, the low intensity cells had increased type I procollagen mRNAs while the high
intensity cells had increased type III procollagen mRNA. The subpopulations of rat lung fibroblasts after isolation using
the fluorescent cell sorter were readily propagated for at least four passages.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1016/S0021-9258(19)39323-8</identifier><identifier>PMID: 2180946</identifier><identifier>CODEN: JBCHA3</identifier><language>eng</language><publisher>Bethesda, MD: American Society for Biochemistry and Molecular Biology</publisher><subject>Animal cells ; Animals ; Antibodies ; Biological and medical sciences ; Cell cultures. Hybridization. Fusion ; Collagen - genetics ; DNA, Recombinant - metabolism ; Fibroblasts - cytology ; Fibroblasts - metabolism ; Flow Cytometry ; Fluorescent Antibody Technique ; Fundamental and applied biological sciences. Psychology ; Lung - cytology ; Lung - metabolism ; Male ; Molecular and cellular biology ; Plasmids ; Procollagen - genetics ; Rats ; Rats, Inbred F344 ; RNA, Messenger - analysis ; RNA, Messenger - genetics</subject><ispartof>The Journal of biological chemistry, 1990-04, Vol.265 (11), p.6286-6290</ispartof><rights>1991 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3248-12ec873fafef0e10a03f2ac91daa5f1f81a025742931adcea830b9915a80db013</citedby><cites>FETCH-LOGICAL-c3248-12ec873fafef0e10a03f2ac91daa5f1f81a025742931adcea830b9915a80db013</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=19251106$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2180946$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>BREEN, E</creatorcontrib><creatorcontrib>FALCO, V. M</creatorcontrib><creatorcontrib>ABSHER, M</creatorcontrib><creatorcontrib>CUTRONEO, K. R</creatorcontrib><title>Subpopulations of rat lung fibroblasts with different amounts of type I and type III collagen mRNAs</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>A fluorescence-based method using the cell sorter has been devised to separate rat lung fibroblasts into subpopulations. Type
I or type III collagen antiserum was used as the primary antibody to react with parent rat lung fibroblasts. This was followed
by a fluorescein-conjugated secondary antibody. Specificity of the primary collagen antibody was determined using a monoclonal
beta-actin antibody and purified IgG as the primary antibodies. The fluorescent shift of parent rat lung fibroblasts was optimized
for the amount of primary collagen antibody and secondary fluorescein-conjugated antibody. An increase in slot blot intensity
was observed for pro-alpha 1(I), pro-alpha 2(I), and pro-alpha 1(III) mRNAs with increasing amounts of cellular RNA. When
precipitating with type I collagen antibodies, the total cellular steady-state levels of type I procollagen mRNAs were increased
in the high intensity cells as compared with the low intensity cells. Alternately, when the type III collagen antibodies were
used to precipitate the rat lung fibroblasts, the low intensity cells had increased type I procollagen mRNAs while the high
intensity cells had increased type III procollagen mRNA. The subpopulations of rat lung fibroblasts after isolation using
the fluorescent cell sorter were readily propagated for at least four passages.</description><subject>Animal cells</subject><subject>Animals</subject><subject>Antibodies</subject><subject>Biological and medical sciences</subject><subject>Cell cultures. Hybridization. Fusion</subject><subject>Collagen - genetics</subject><subject>DNA, Recombinant - metabolism</subject><subject>Fibroblasts - cytology</subject><subject>Fibroblasts - metabolism</subject><subject>Flow Cytometry</subject><subject>Fluorescent Antibody Technique</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Lung - cytology</subject><subject>Lung - metabolism</subject><subject>Male</subject><subject>Molecular and cellular biology</subject><subject>Plasmids</subject><subject>Procollagen - genetics</subject><subject>Rats</subject><subject>Rats, Inbred F344</subject><subject>RNA, Messenger - analysis</subject><subject>RNA, Messenger - genetics</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1990</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpFkN1q3DAQRkVJSLdJHyEgKAnthRuNtLalyxDSdmFpIT-QOzGWpV0V23Ilm5C3r3fXbHQzgjnzzXAIuQT2HRgUN4-MccgUz-VXUN-EElxk8gNZAJMiEzm8nJDFEflIPqX0l01vqeCMnHGQTC2LBTGPY9WHfmxw8KFLNDgacaDN2G2o81UMVYNpSPTVD1tae-dstN1AsQ1jN-zx4a23dEWxq-fvakVNaBrc2I62D79v0wU5ddgk-3mu5-T5x_3T3a9s_efn6u52nRnBlzIDbo0shUNnHbPAkAnH0SioEXMHTgIynpdLrgRgbSxKwSqlIEfJ6oqBOCfXh9w-hn-jTYNufTJ2OqWzYUwa8hJ4WZQTmB9AE0NK0TrdR99ifNPA9E6u3svVO3MalN7L1XKau5wXjFVr6-PUbHPqX819TAYbF7EzPr2HT3kAbMd9OXBbv9m--mh15YPZ2lbzItcAuuCyEP8B0g6N1A</recordid><startdate>19900415</startdate><enddate>19900415</enddate><creator>BREEN, E</creator><creator>FALCO, V. M</creator><creator>ABSHER, M</creator><creator>CUTRONEO, K. R</creator><general>American Society for Biochemistry and Molecular Biology</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>7TM</scope></search><sort><creationdate>19900415</creationdate><title>Subpopulations of rat lung fibroblasts with different amounts of type I and type III collagen mRNAs</title><author>BREEN, E ; FALCO, V. M ; ABSHER, M ; CUTRONEO, K. R</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3248-12ec873fafef0e10a03f2ac91daa5f1f81a025742931adcea830b9915a80db013</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1990</creationdate><topic>Animal cells</topic><topic>Animals</topic><topic>Antibodies</topic><topic>Biological and medical sciences</topic><topic>Cell cultures. Hybridization. Fusion</topic><topic>Collagen - genetics</topic><topic>DNA, Recombinant - metabolism</topic><topic>Fibroblasts - cytology</topic><topic>Fibroblasts - metabolism</topic><topic>Flow Cytometry</topic><topic>Fluorescent Antibody Technique</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Lung - cytology</topic><topic>Lung - metabolism</topic><topic>Male</topic><topic>Molecular and cellular biology</topic><topic>Plasmids</topic><topic>Procollagen - genetics</topic><topic>Rats</topic><topic>Rats, Inbred F344</topic><topic>RNA, Messenger - analysis</topic><topic>RNA, Messenger - genetics</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>BREEN, E</creatorcontrib><creatorcontrib>FALCO, V. M</creatorcontrib><creatorcontrib>ABSHER, M</creatorcontrib><creatorcontrib>CUTRONEO, K. R</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>BREEN, E</au><au>FALCO, V. M</au><au>ABSHER, M</au><au>CUTRONEO, K. R</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Subpopulations of rat lung fibroblasts with different amounts of type I and type III collagen mRNAs</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>1990-04-15</date><risdate>1990</risdate><volume>265</volume><issue>11</issue><spage>6286</spage><epage>6290</epage><pages>6286-6290</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><coden>JBCHA3</coden><abstract>A fluorescence-based method using the cell sorter has been devised to separate rat lung fibroblasts into subpopulations. Type
I or type III collagen antiserum was used as the primary antibody to react with parent rat lung fibroblasts. This was followed
by a fluorescein-conjugated secondary antibody. Specificity of the primary collagen antibody was determined using a monoclonal
beta-actin antibody and purified IgG as the primary antibodies. The fluorescent shift of parent rat lung fibroblasts was optimized
for the amount of primary collagen antibody and secondary fluorescein-conjugated antibody. An increase in slot blot intensity
was observed for pro-alpha 1(I), pro-alpha 2(I), and pro-alpha 1(III) mRNAs with increasing amounts of cellular RNA. When
precipitating with type I collagen antibodies, the total cellular steady-state levels of type I procollagen mRNAs were increased
in the high intensity cells as compared with the low intensity cells. Alternately, when the type III collagen antibodies were
used to precipitate the rat lung fibroblasts, the low intensity cells had increased type I procollagen mRNAs while the high
intensity cells had increased type III procollagen mRNA. The subpopulations of rat lung fibroblasts after isolation using
the fluorescent cell sorter were readily propagated for at least four passages.</abstract><cop>Bethesda, MD</cop><pub>American Society for Biochemistry and Molecular Biology</pub><pmid>2180946</pmid><doi>10.1016/S0021-9258(19)39323-8</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | The Journal of biological chemistry, 1990-04, Vol.265 (11), p.6286-6290 |
| issn | 0021-9258 1083-351X |
| language | eng |
| recordid | cdi_proquest_miscellaneous_15712767 |
| source | MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection |
| subjects | Animal cells Animals Antibodies Biological and medical sciences Cell cultures. Hybridization. Fusion Collagen - genetics DNA, Recombinant - metabolism Fibroblasts - cytology Fibroblasts - metabolism Flow Cytometry Fluorescent Antibody Technique Fundamental and applied biological sciences. Psychology Lung - cytology Lung - metabolism Male Molecular and cellular biology Plasmids Procollagen - genetics Rats Rats, Inbred F344 RNA, Messenger - analysis RNA, Messenger - genetics |
| title | Subpopulations of rat lung fibroblasts with different amounts of type I and type III collagen mRNAs |
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