Optimized large-scale production of high titer lentivirus vector pseudotypes
The goal of the present study was to develop an efficient transient transfection method for large-scale production of high titer lentivirus vector stocks of eight different pseudotypes. The envelope genes used for this purpose were those from VSV-G, Mokola, Rabies, MLV-Ampho, MLV-10A1, LCMV-WE, and...
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| Veröffentlicht in: | Journal of virological methods 2004-12, Vol.122 (2), p.131-139 |
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| creator | Sena-Esteves, Miguel Tebbets, Jessica C. Steffens, Sabine Crombleholme, Timothy Flake, Alan W. |
| description | The goal of the present study was to develop an efficient transient transfection method for large-scale production of high titer lentivirus vector stocks of eight different pseudotypes. The envelope genes used for this purpose were those from VSV-G, Mokola, Rabies, MLV-Ampho, MLV-10A1, LCMV-WE, and LCMV-Arm53b. All envelopes were cloned into phCMV, which yielded lentivirus vector titers one, two, or three orders of magnitude higher than the original plasmids for the Rabies, MLV-10A1, and MLV-Ampho envelopes, respectively. When these newly constructed envelope expression plasmids were used for packaging, treatment with sodium butyrate resulted in almost five-fold increase in titers for some of the pseudotypes, had no effect for others (VSV-G and Rabies), and negatively impacted titers for the LCMV-derived pseudotypes. Production of vectors in serum-free media yielded titers only slightly lower than those obtained in the presence of serum. The efficiency of concentrating vector supernatants by ultracentrifugation or ultrafiltration was compared, with higher recovery efficiencies for the latter method, but the highest titers for most pseudotypes were obtained by ultracentrifugation. The best conditions for each individual pseudotype yielded lentivirus vector stocks with titers above 1
×
10
9
tu/mL for most pseudotypes, and higher than 1
×
10
10
tu/mL for VSV-G. |
| doi_str_mv | 10.1016/j.jviromet.2004.08.017 |
| format | Article |
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×
10
9
tu/mL for most pseudotypes, and higher than 1
×
10
10
tu/mL for VSV-G.</description><identifier>ISSN: 0166-0934</identifier><identifier>EISSN: 1879-0984</identifier><identifier>DOI: 10.1016/j.jviromet.2004.08.017</identifier><identifier>PMID: 15542136</identifier><identifier>CODEN: JVMEDH</identifier><language>eng</language><publisher>London: Elsevier B.V</publisher><subject>10A1 ; Amphotropic ; Biological and medical sciences ; Cell Line ; Ebolavirus - genetics ; Fundamental and applied biological sciences. Psychology ; Genetic Vectors - genetics ; Genetic Vectors - physiology ; HIV-1 - genetics ; HIV-1 - physiology ; LCMV ; Lentivirus ; Lentivirus - genetics ; Lentivirus - physiology ; Lymphocytic choriomeningitis virus - genetics ; Microbiology ; Mokola ; Plasmids ; Pseudotype ; Rabies ; Techniques used in virology ; Transduction, Genetic - methods ; Transfection ; Vector ; Vesicular stomatitis Indiana virus - genetics ; Viral Envelope Proteins - genetics ; Viral Fusion Proteins - genetics ; Virology ; VSV-G</subject><ispartof>Journal of virological methods, 2004-12, Vol.122 (2), p.131-139</ispartof><rights>2004 Elsevier B.V.</rights><rights>2005 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c487t-c4084343da9e781a4c30c28123a6ca71888a711b280b02fcd6cfa4428e9df1313</citedby><cites>FETCH-LOGICAL-c487t-c4084343da9e781a4c30c28123a6ca71888a711b280b02fcd6cfa4428e9df1313</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.jviromet.2004.08.017$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3541,27915,27916,45986</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=16291809$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15542136$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Sena-Esteves, Miguel</creatorcontrib><creatorcontrib>Tebbets, Jessica C.</creatorcontrib><creatorcontrib>Steffens, Sabine</creatorcontrib><creatorcontrib>Crombleholme, Timothy</creatorcontrib><creatorcontrib>Flake, Alan W.</creatorcontrib><title>Optimized large-scale production of high titer lentivirus vector pseudotypes</title><title>Journal of virological methods</title><addtitle>J Virol Methods</addtitle><description>The goal of the present study was to develop an efficient transient transfection method for large-scale production of high titer lentivirus vector stocks of eight different pseudotypes. The envelope genes used for this purpose were those from VSV-G, Mokola, Rabies, MLV-Ampho, MLV-10A1, LCMV-WE, and LCMV-Arm53b. All envelopes were cloned into phCMV, which yielded lentivirus vector titers one, two, or three orders of magnitude higher than the original plasmids for the Rabies, MLV-10A1, and MLV-Ampho envelopes, respectively. When these newly constructed envelope expression plasmids were used for packaging, treatment with sodium butyrate resulted in almost five-fold increase in titers for some of the pseudotypes, had no effect for others (VSV-G and Rabies), and negatively impacted titers for the LCMV-derived pseudotypes. Production of vectors in serum-free media yielded titers only slightly lower than those obtained in the presence of serum. The efficiency of concentrating vector supernatants by ultracentrifugation or ultrafiltration was compared, with higher recovery efficiencies for the latter method, but the highest titers for most pseudotypes were obtained by ultracentrifugation. The best conditions for each individual pseudotype yielded lentivirus vector stocks with titers above 1
×
10
9
tu/mL for most pseudotypes, and higher than 1
×
10
10
tu/mL for VSV-G.</description><subject>10A1</subject><subject>Amphotropic</subject><subject>Biological and medical sciences</subject><subject>Cell Line</subject><subject>Ebolavirus - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Genetic Vectors - genetics</subject><subject>Genetic Vectors - physiology</subject><subject>HIV-1 - genetics</subject><subject>HIV-1 - physiology</subject><subject>LCMV</subject><subject>Lentivirus</subject><subject>Lentivirus - genetics</subject><subject>Lentivirus - physiology</subject><subject>Lymphocytic choriomeningitis virus - genetics</subject><subject>Microbiology</subject><subject>Mokola</subject><subject>Plasmids</subject><subject>Pseudotype</subject><subject>Rabies</subject><subject>Techniques used in virology</subject><subject>Transduction, Genetic - methods</subject><subject>Transfection</subject><subject>Vector</subject><subject>Vesicular stomatitis Indiana virus - genetics</subject><subject>Viral Envelope Proteins - genetics</subject><subject>Viral Fusion Proteins - genetics</subject><subject>Virology</subject><subject>VSV-G</subject><issn>0166-0934</issn><issn>1879-0984</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNkTtv2zAQgImgReOk_QuBlnazenxIpLYWQZMWMJClnQmaPCU0JFEhKQPprw8Nu8joLnc3fPfCR8gNhZoCbb_u6t3exzBirhmAqEHVQOUFWVEluzV0SrwjqwK2pebiklyltAOARnL-gVzSphGM8nZFNg9z9qP_i64aTHzEdbJmwGqOwS02-zBVoa-e_ONTlX3GWA04ZV82L6nao80hVnPCxYX8MmP6SN73Zkj46ZSvyZ-7H79vf643D_e_br9v1lYomUsEJbjgznQoFTXCcrBMUcZNa42kSqkS6ZYp2ALrrWttb4RgCjvXU075NflynFvOfF4wZT36ZHEYzIRhSbqV0ErB-VmQdQo6KeV_gA3nDRNnQSqlkpSzArZH0MaQUsRez9GPJr5oCvqgUO_0P4X6oFCD0kVhabw5bVi2I7q3tpOzAnw-AeYgq49msj69cS3raHmrcN-OHBYVe49RJ-txsuh8LOq0C_7cLa8WKb2B</recordid><startdate>20041215</startdate><enddate>20041215</enddate><creator>Sena-Esteves, Miguel</creator><creator>Tebbets, Jessica C.</creator><creator>Steffens, Sabine</creator><creator>Crombleholme, Timothy</creator><creator>Flake, Alan W.</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>7T7</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>P64</scope><scope>7TB</scope><scope>F28</scope><scope>7X8</scope></search><sort><creationdate>20041215</creationdate><title>Optimized large-scale production of high titer lentivirus vector pseudotypes</title><author>Sena-Esteves, Miguel ; Tebbets, Jessica C. ; Steffens, Sabine ; Crombleholme, Timothy ; Flake, Alan W.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c487t-c4084343da9e781a4c30c28123a6ca71888a711b280b02fcd6cfa4428e9df1313</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2004</creationdate><topic>10A1</topic><topic>Amphotropic</topic><topic>Biological and medical sciences</topic><topic>Cell Line</topic><topic>Ebolavirus - genetics</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Genetic Vectors - genetics</topic><topic>Genetic Vectors - physiology</topic><topic>HIV-1 - genetics</topic><topic>HIV-1 - physiology</topic><topic>LCMV</topic><topic>Lentivirus</topic><topic>Lentivirus - genetics</topic><topic>Lentivirus - physiology</topic><topic>Lymphocytic choriomeningitis virus - genetics</topic><topic>Microbiology</topic><topic>Mokola</topic><topic>Plasmids</topic><topic>Pseudotype</topic><topic>Rabies</topic><topic>Techniques used in virology</topic><topic>Transduction, Genetic - methods</topic><topic>Transfection</topic><topic>Vector</topic><topic>Vesicular stomatitis Indiana virus - genetics</topic><topic>Viral Envelope Proteins - genetics</topic><topic>Viral Fusion Proteins - genetics</topic><topic>Virology</topic><topic>VSV-G</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Sena-Esteves, Miguel</creatorcontrib><creatorcontrib>Tebbets, Jessica C.</creatorcontrib><creatorcontrib>Steffens, Sabine</creatorcontrib><creatorcontrib>Crombleholme, Timothy</creatorcontrib><creatorcontrib>Flake, Alan W.</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>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Mechanical & Transportation Engineering Abstracts</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of virological methods</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Sena-Esteves, Miguel</au><au>Tebbets, Jessica C.</au><au>Steffens, Sabine</au><au>Crombleholme, Timothy</au><au>Flake, Alan W.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Optimized large-scale production of high titer lentivirus vector pseudotypes</atitle><jtitle>Journal of virological methods</jtitle><addtitle>J Virol Methods</addtitle><date>2004-12-15</date><risdate>2004</risdate><volume>122</volume><issue>2</issue><spage>131</spage><epage>139</epage><pages>131-139</pages><issn>0166-0934</issn><eissn>1879-0984</eissn><coden>JVMEDH</coden><abstract>The goal of the present study was to develop an efficient transient transfection method for large-scale production of high titer lentivirus vector stocks of eight different pseudotypes. The envelope genes used for this purpose were those from VSV-G, Mokola, Rabies, MLV-Ampho, MLV-10A1, LCMV-WE, and LCMV-Arm53b. All envelopes were cloned into phCMV, which yielded lentivirus vector titers one, two, or three orders of magnitude higher than the original plasmids for the Rabies, MLV-10A1, and MLV-Ampho envelopes, respectively. When these newly constructed envelope expression plasmids were used for packaging, treatment with sodium butyrate resulted in almost five-fold increase in titers for some of the pseudotypes, had no effect for others (VSV-G and Rabies), and negatively impacted titers for the LCMV-derived pseudotypes. Production of vectors in serum-free media yielded titers only slightly lower than those obtained in the presence of serum. The efficiency of concentrating vector supernatants by ultracentrifugation or ultrafiltration was compared, with higher recovery efficiencies for the latter method, but the highest titers for most pseudotypes were obtained by ultracentrifugation. The best conditions for each individual pseudotype yielded lentivirus vector stocks with titers above 1
×
10
9
tu/mL for most pseudotypes, and higher than 1
×
10
10
tu/mL for VSV-G.</abstract><cop>London</cop><cop>Amsterdam</cop><cop>New York, NY</cop><pub>Elsevier B.V</pub><pmid>15542136</pmid><doi>10.1016/j.jviromet.2004.08.017</doi><tpages>9</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0166-0934 |
| ispartof | Journal of virological methods, 2004-12, Vol.122 (2), p.131-139 |
| issn | 0166-0934 1879-0984 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_67067433 |
| source | MEDLINE; ScienceDirect Journals (5 years ago - present) |
| subjects | 10A1 Amphotropic Biological and medical sciences Cell Line Ebolavirus - genetics Fundamental and applied biological sciences. Psychology Genetic Vectors - genetics Genetic Vectors - physiology HIV-1 - genetics HIV-1 - physiology LCMV Lentivirus Lentivirus - genetics Lentivirus - physiology Lymphocytic choriomeningitis virus - genetics Microbiology Mokola Plasmids Pseudotype Rabies Techniques used in virology Transduction, Genetic - methods Transfection Vector Vesicular stomatitis Indiana virus - genetics Viral Envelope Proteins - genetics Viral Fusion Proteins - genetics Virology VSV-G |
| title | Optimized large-scale production of high titer lentivirus vector pseudotypes |
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