Optimizing follicular dendritic cell isolation by discontinuous gradient centrifugation and use of the magnetic cell sorter (MACS)
Follicular dendritic cells (FDC) contribute minimally to the total cell population of lymphatic tissue. In order to obtain higher numbers of viable FDC with only a small fraction of contaminating cells the following procedure was developed. Subsequent to the usual mechanical and enzymatical digestio...
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| Veröffentlicht in: | Journal of immunological methods 1993-02, Vol.159 (1), p.189-196 |
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| container_title | Journal of immunological methods |
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| creator | Schmitz, Jörn Petrasch, Stephan van Lunzen, Jan Racz, Paul Kleine, Hans-Dieter Hufert, Frank Kern, Peter Schmitz, Herbert Tenner-Racz, Klara |
| description | Follicular dendritic cells (FDC) contribute minimally to the total cell population of lymphatic tissue. In order to obtain higher numbers of viable FDC with only a small fraction of contaminating cells the following procedure was developed. Subsequent to the usual mechanical and enzymatical digestion of human tonsils, single cells were layered on top of a discontinuous bovine albumin gradient and centrifuged at 8500 ×
g. The suspension collected from the 1.052-1.030 interphase contained an average of 10.5% FDC. Next, the preparation was subjected to a new step involving separation of FDC previously treated with biotin-labelled KiM4 monoclonal antibody, raised against FDC, and attached via biotin-streptavidin bonding to streptavidin-conjugated paramagnetic beads. Purification on a magnetic cell sorter (MACS) yielded 3.3−10.1×10
6 cells with an average FDC content of 78.4%. The viability and morphology of the resulting FDC population was examined using trypan blue staining or electron microscopy. This technique will permit in vitro studies and long term cultures with FDC isolated from human lymphatic tissue. |
| doi_str_mv | 10.1016/0022-1759(93)90157-3 |
| format | Article |
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g. The suspension collected from the 1.052-1.030 interphase contained an average of 10.5% FDC. Next, the preparation was subjected to a new step involving separation of FDC previously treated with biotin-labelled KiM4 monoclonal antibody, raised against FDC, and attached via biotin-streptavidin bonding to streptavidin-conjugated paramagnetic beads. Purification on a magnetic cell sorter (MACS) yielded 3.3−10.1×10
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g. The suspension collected from the 1.052-1.030 interphase contained an average of 10.5% FDC. Next, the preparation was subjected to a new step involving separation of FDC previously treated with biotin-labelled KiM4 monoclonal antibody, raised against FDC, and attached via biotin-streptavidin bonding to streptavidin-conjugated paramagnetic beads. Purification on a magnetic cell sorter (MACS) yielded 3.3−10.1×10
6 cells with an average FDC content of 78.4%. The viability and morphology of the resulting FDC population was examined using trypan blue staining or electron microscopy. This technique will permit in vitro studies and long term cultures with FDC isolated from human lymphatic tissue.</description><subject>Antibodies, Monoclonal - immunology</subject><subject>Biological and medical sciences</subject><subject>Cell Separation - methods</subject><subject>Centrifugation, Density Gradient</subject><subject>Dendritic Cells - cytology</subject><subject>Dendritic Cells - ultrastructure</subject><subject>Follicular dendritic cell</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Fundamental immunology</subject><subject>Humans</subject><subject>Magnetic cell sorter</subject><subject>Magnetics</subject><subject>Microscopy, Electron</subject><subject>Molecular immunology</subject><subject>Techniques</subject><issn>0022-1759</issn><issn>1872-7905</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1993</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkT1vFDEQhi0ECpfAPwDJBUJJseBv3zaRolP4kIJSALXltceH0a592F6kUPLL2eNOV0I1xTzzamYehF5Q8oYSqt4SwlhHtewve37VEyp1xx-hFV1r1umeyMdodUKeovNavxNCKFHkDJ2thZBMshX6fb9rcYq_YtrikMcxunm0BXtIvsQWHXYwjjjWPNoWc8LDA_axupxaTHOeK94W6yOktoCplRjm7QG0yeO5As4Bt2-AJ7tNcMqruTQo-PLTzebz1TP0JNixwvNjvUBf391-2Xzo7u7ff9zc3HVOrHnrgPKgAgwMuKSgFLXae2lBOKcFAw1hUANfK0mI5J455wN3NgyS08HrXvIL9PqQuyv5xwy1mWm5ZFnHJlguMVoqRrgk_wWpEj3VjC6gOICu5FoLBLMrcbLlwVBi9o7MXoDZCzA9N38dGb6MvTzmz8ME_jR0lLL0Xx37tjo7hmKTi_WECSU0UXvs-oDB8rSfEYqpbjHhwMcCrhmf47_3-AOuUa_R</recordid><startdate>19930226</startdate><enddate>19930226</enddate><creator>Schmitz, Jörn</creator><creator>Petrasch, Stephan</creator><creator>van Lunzen, Jan</creator><creator>Racz, Paul</creator><creator>Kleine, Hans-Dieter</creator><creator>Hufert, Frank</creator><creator>Kern, Peter</creator><creator>Schmitz, Herbert</creator><creator>Tenner-Racz, Klara</creator><general>Elsevier B.V</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>7T5</scope><scope>H94</scope><scope>7X8</scope></search><sort><creationdate>19930226</creationdate><title>Optimizing follicular dendritic cell isolation by discontinuous gradient centrifugation and use of the magnetic cell sorter (MACS)</title><author>Schmitz, Jörn ; Petrasch, Stephan ; van Lunzen, Jan ; Racz, Paul ; Kleine, Hans-Dieter ; Hufert, Frank ; Kern, Peter ; Schmitz, Herbert ; Tenner-Racz, Klara</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c483t-e13f6feb2e351e661a7dd5ae4cc742e7efb6b38650053d2ccdf3cafb531bd7953</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1993</creationdate><topic>Antibodies, Monoclonal - immunology</topic><topic>Biological and medical sciences</topic><topic>Cell Separation - methods</topic><topic>Centrifugation, Density Gradient</topic><topic>Dendritic Cells - cytology</topic><topic>Dendritic Cells - ultrastructure</topic><topic>Follicular dendritic cell</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Fundamental immunology</topic><topic>Humans</topic><topic>Magnetic cell sorter</topic><topic>Magnetics</topic><topic>Microscopy, Electron</topic><topic>Molecular immunology</topic><topic>Techniques</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Schmitz, Jörn</creatorcontrib><creatorcontrib>Petrasch, Stephan</creatorcontrib><creatorcontrib>van Lunzen, Jan</creatorcontrib><creatorcontrib>Racz, Paul</creatorcontrib><creatorcontrib>Kleine, Hans-Dieter</creatorcontrib><creatorcontrib>Hufert, Frank</creatorcontrib><creatorcontrib>Kern, Peter</creatorcontrib><creatorcontrib>Schmitz, Herbert</creatorcontrib><creatorcontrib>Tenner-Racz, Klara</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>Immunology Abstracts</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of immunological methods</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Schmitz, Jörn</au><au>Petrasch, Stephan</au><au>van Lunzen, Jan</au><au>Racz, Paul</au><au>Kleine, Hans-Dieter</au><au>Hufert, Frank</au><au>Kern, Peter</au><au>Schmitz, Herbert</au><au>Tenner-Racz, Klara</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Optimizing follicular dendritic cell isolation by discontinuous gradient centrifugation and use of the magnetic cell sorter (MACS)</atitle><jtitle>Journal of immunological methods</jtitle><addtitle>J Immunol Methods</addtitle><date>1993-02-26</date><risdate>1993</risdate><volume>159</volume><issue>1</issue><spage>189</spage><epage>196</epage><pages>189-196</pages><issn>0022-1759</issn><eissn>1872-7905</eissn><coden>JIMMBG</coden><abstract>Follicular dendritic cells (FDC) contribute minimally to the total cell population of lymphatic tissue. In order to obtain higher numbers of viable FDC with only a small fraction of contaminating cells the following procedure was developed. Subsequent to the usual mechanical and enzymatical digestion of human tonsils, single cells were layered on top of a discontinuous bovine albumin gradient and centrifuged at 8500 ×
g. The suspension collected from the 1.052-1.030 interphase contained an average of 10.5% FDC. Next, the preparation was subjected to a new step involving separation of FDC previously treated with biotin-labelled KiM4 monoclonal antibody, raised against FDC, and attached via biotin-streptavidin bonding to streptavidin-conjugated paramagnetic beads. Purification on a magnetic cell sorter (MACS) yielded 3.3−10.1×10
6 cells with an average FDC content of 78.4%. The viability and morphology of the resulting FDC population was examined using trypan blue staining or electron microscopy. This technique will permit in vitro studies and long term cultures with FDC isolated from human lymphatic tissue.</abstract><cop>Amsterdam</cop><pub>Elsevier B.V</pub><pmid>8445252</pmid><doi>10.1016/0022-1759(93)90157-3</doi><tpages>8</tpages></addata></record> |
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| subjects | Antibodies, Monoclonal - immunology Biological and medical sciences Cell Separation - methods Centrifugation, Density Gradient Dendritic Cells - cytology Dendritic Cells - ultrastructure Follicular dendritic cell Fundamental and applied biological sciences. Psychology Fundamental immunology Humans Magnetic cell sorter Magnetics Microscopy, Electron Molecular immunology Techniques |
| title | Optimizing follicular dendritic cell isolation by discontinuous gradient centrifugation and use of the magnetic cell sorter (MACS) |
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