Characterization and downstream processing of HIV-1 core and virus-like-particles produced in serum free medium
This work aimed at the characterisation and downstream processing of HIV-1 core and virus-like particles (CLPs and VLPs) produced in serum free medium. The produced core and virus-like particles have similar characteristics to those of the native immature HIV-1 virus in terms of protein composition,...
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| Veröffentlicht in: | Enzyme and microbial technology 2000, Vol.26 (1), p.61-70 |
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| creator | Cruz, Pedro E Peixoto, Cristina C Devos, Kathleen Moreira, José L Saman, Eric Carrondo, Manuel J.T |
| description | This work aimed at the characterisation and downstream processing of HIV-1 core and virus-like particles (CLPs and VLPs) produced in serum free medium. The produced core and virus-like particles have similar characteristics to those of the native immature HIV-1 virus in terms of protein composition, diameter, density and shape. The molecular mass (determined by gel filtration chromatography) and the diameter (determined by electron microscopy) were similar for CLPs and VLPs, this being correlated with the difference between the particle densities (1.14 g/cm
3 and 1.19 g/cm
3, respectively, for CLPs and VLPs). After this characterization, several concentration and purification methods were tested to obtain enough material for further tests. For this purpose, a downstream processing strategy was developed, involving clarification by centrifugation and microfiltration, concentration and partial purification by ultrafiltration, and final purification by gel filtration chromatography. Quality analysis using Western immunoblot and gel filtration chromatography was performed to define the best operational conditions for each purification step. The particle recovery obtained was 87% with the main losses occurring in the ultrafiltration step. In addition, by evaluating the effect of operational conditions on product quality, it was possible to obtain a final product with high content of intact high molecular weight particles. |
| doi_str_mv | 10.1016/S0141-0229(99)00128-3 |
| format | Article |
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3 and 1.19 g/cm
3, respectively, for CLPs and VLPs). After this characterization, several concentration and purification methods were tested to obtain enough material for further tests. For this purpose, a downstream processing strategy was developed, involving clarification by centrifugation and microfiltration, concentration and partial purification by ultrafiltration, and final purification by gel filtration chromatography. Quality analysis using Western immunoblot and gel filtration chromatography was performed to define the best operational conditions for each purification step. The particle recovery obtained was 87% with the main losses occurring in the ultrafiltration step. In addition, by evaluating the effect of operational conditions on product quality, it was possible to obtain a final product with high content of intact high molecular weight particles.</description><identifier>ISSN: 0141-0229</identifier><identifier>EISSN: 1879-0909</identifier><identifier>DOI: 10.1016/S0141-0229(99)00128-3</identifier><identifier>CODEN: EMTED2</identifier><language>eng</language><publisher>Amsterdam: Elsevier Inc</publisher><subject>Biological and medical sciences ; Biotechnology ; Centrifugation ; Characterization ; Chromatography ; Clarification ; Density (specific gravity) ; Fundamental and applied biological sciences. Psychology ; Health. Pharmaceutical industry ; HIV-1 ; Human immunodeficiency virus 1 ; Industrial applications and implications. Economical aspects ; Insect cells ; Microfiltration ; Molecular weight ; Other active biomolecules ; Production of active biomolecules ; Purification ; Ultrafiltration ; Virus-like particles ; Viruses</subject><ispartof>Enzyme and microbial technology, 2000, Vol.26 (1), p.61-70</ispartof><rights>2000 Elsevier Science Inc.</rights><rights>2000 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c398t-23eb2707d2895c4fa55c2209db4dac06fd3357c09011d8ae3a16dda0f8b6f9233</citedby><cites>FETCH-LOGICAL-c398t-23eb2707d2895c4fa55c2209db4dac06fd3357c09011d8ae3a16dda0f8b6f9233</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0141022999001283$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,776,780,3536,4009,27902,27903,27904,65309</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=1275968$$DView record in Pascal Francis$$Hfree_for_read</backlink></links><search><creatorcontrib>Cruz, Pedro E</creatorcontrib><creatorcontrib>Peixoto, Cristina C</creatorcontrib><creatorcontrib>Devos, Kathleen</creatorcontrib><creatorcontrib>Moreira, José L</creatorcontrib><creatorcontrib>Saman, Eric</creatorcontrib><creatorcontrib>Carrondo, Manuel J.T</creatorcontrib><title>Characterization and downstream processing of HIV-1 core and virus-like-particles produced in serum free medium</title><title>Enzyme and microbial technology</title><description>This work aimed at the characterisation and downstream processing of HIV-1 core and virus-like particles (CLPs and VLPs) produced in serum free medium. The produced core and virus-like particles have similar characteristics to those of the native immature HIV-1 virus in terms of protein composition, diameter, density and shape. The molecular mass (determined by gel filtration chromatography) and the diameter (determined by electron microscopy) were similar for CLPs and VLPs, this being correlated with the difference between the particle densities (1.14 g/cm
3 and 1.19 g/cm
3, respectively, for CLPs and VLPs). After this characterization, several concentration and purification methods were tested to obtain enough material for further tests. For this purpose, a downstream processing strategy was developed, involving clarification by centrifugation and microfiltration, concentration and partial purification by ultrafiltration, and final purification by gel filtration chromatography. Quality analysis using Western immunoblot and gel filtration chromatography was performed to define the best operational conditions for each purification step. The particle recovery obtained was 87% with the main losses occurring in the ultrafiltration step. In addition, by evaluating the effect of operational conditions on product quality, it was possible to obtain a final product with high content of intact high molecular weight particles.</description><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Centrifugation</subject><subject>Characterization</subject><subject>Chromatography</subject><subject>Clarification</subject><subject>Density (specific gravity)</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Health. Pharmaceutical industry</subject><subject>HIV-1</subject><subject>Human immunodeficiency virus 1</subject><subject>Industrial applications and implications. Economical aspects</subject><subject>Insect cells</subject><subject>Microfiltration</subject><subject>Molecular weight</subject><subject>Other active biomolecules</subject><subject>Production of active biomolecules</subject><subject>Purification</subject><subject>Ultrafiltration</subject><subject>Virus-like particles</subject><subject>Viruses</subject><issn>0141-0229</issn><issn>1879-0909</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2000</creationdate><recordtype>article</recordtype><recordid>eNqFkUtvFDEQhC0EEkvIT0DyAaFwcPBjXj5F0SqQSJE4QHK1eu02GGbGi3smiPx6Znej5JhTX77uqq5i7J2Sp0qq5tM3qSolpNb2xNqPUirdCfOCrVTXWiGttC_Z6hF5zd4Q_ZILVVVyxfL6JxTwE5Z0D1PKI4cx8JD_jjQVhIFvS_ZIlMYfPEd-eXUrFPe54J67S2Um0affKLZQpuR7pN1GmD0GnkZOWOaBx4LIBwxpHt6yVxF6wuOHecRuPl98X1-K669frtbn18Ib201CG9zoVrZBd7b2VYS69lpLGzZVAC-bGIypW788p1ToAA2oJgSQsds00WpjjtiHw93FzZ8ZaXJDIo99DyPmmZxWpmsqZZ8FVVsr1bRyAesD6EsmKhjdtqQByj-npNv14PY9uF3Izlq378HtnLx_EADy0McCo0_0tKzb2jbdgp0dMFxSuUtYHPmE45JjKugnF3J6Rug_JCKdLA</recordid><startdate>2000</startdate><enddate>2000</enddate><creator>Cruz, Pedro E</creator><creator>Peixoto, Cristina C</creator><creator>Devos, Kathleen</creator><creator>Moreira, José L</creator><creator>Saman, Eric</creator><creator>Carrondo, Manuel J.T</creator><general>Elsevier Inc</general><general>Elsevier Science</general><scope>IQODW</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7U9</scope><scope>H94</scope></search><sort><creationdate>2000</creationdate><title>Characterization and downstream processing of HIV-1 core and virus-like-particles produced in serum free medium</title><author>Cruz, Pedro E ; Peixoto, Cristina C ; Devos, Kathleen ; Moreira, José L ; Saman, Eric ; Carrondo, Manuel J.T</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c398t-23eb2707d2895c4fa55c2209db4dac06fd3357c09011d8ae3a16dda0f8b6f9233</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2000</creationdate><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Centrifugation</topic><topic>Characterization</topic><topic>Chromatography</topic><topic>Clarification</topic><topic>Density (specific gravity)</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Health. Pharmaceutical industry</topic><topic>HIV-1</topic><topic>Human immunodeficiency virus 1</topic><topic>Industrial applications and implications. Economical aspects</topic><topic>Insect cells</topic><topic>Microfiltration</topic><topic>Molecular weight</topic><topic>Other active biomolecules</topic><topic>Production of active biomolecules</topic><topic>Purification</topic><topic>Ultrafiltration</topic><topic>Virus-like particles</topic><topic>Viruses</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Cruz, Pedro E</creatorcontrib><creatorcontrib>Peixoto, Cristina C</creatorcontrib><creatorcontrib>Devos, Kathleen</creatorcontrib><creatorcontrib>Moreira, José L</creatorcontrib><creatorcontrib>Saman, Eric</creatorcontrib><creatorcontrib>Carrondo, Manuel J.T</creatorcontrib><collection>Pascal-Francis</collection><collection>CrossRef</collection><collection>Virology and AIDS Abstracts</collection><collection>AIDS and Cancer Research Abstracts</collection><jtitle>Enzyme and microbial technology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Cruz, Pedro E</au><au>Peixoto, Cristina C</au><au>Devos, Kathleen</au><au>Moreira, José L</au><au>Saman, Eric</au><au>Carrondo, Manuel J.T</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Characterization and downstream processing of HIV-1 core and virus-like-particles produced in serum free medium</atitle><jtitle>Enzyme and microbial technology</jtitle><date>2000</date><risdate>2000</risdate><volume>26</volume><issue>1</issue><spage>61</spage><epage>70</epage><pages>61-70</pages><issn>0141-0229</issn><eissn>1879-0909</eissn><coden>EMTED2</coden><abstract>This work aimed at the characterisation and downstream processing of HIV-1 core and virus-like particles (CLPs and VLPs) produced in serum free medium. The produced core and virus-like particles have similar characteristics to those of the native immature HIV-1 virus in terms of protein composition, diameter, density and shape. The molecular mass (determined by gel filtration chromatography) and the diameter (determined by electron microscopy) were similar for CLPs and VLPs, this being correlated with the difference between the particle densities (1.14 g/cm
3 and 1.19 g/cm
3, respectively, for CLPs and VLPs). After this characterization, several concentration and purification methods were tested to obtain enough material for further tests. For this purpose, a downstream processing strategy was developed, involving clarification by centrifugation and microfiltration, concentration and partial purification by ultrafiltration, and final purification by gel filtration chromatography. Quality analysis using Western immunoblot and gel filtration chromatography was performed to define the best operational conditions for each purification step. The particle recovery obtained was 87% with the main losses occurring in the ultrafiltration step. In addition, by evaluating the effect of operational conditions on product quality, it was possible to obtain a final product with high content of intact high molecular weight particles.</abstract><cop>Amsterdam</cop><pub>Elsevier Inc</pub><doi>10.1016/S0141-0229(99)00128-3</doi><tpages>10</tpages></addata></record> |
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| source | Elsevier ScienceDirect Journals |
| subjects | Biological and medical sciences Biotechnology Centrifugation Characterization Chromatography Clarification Density (specific gravity) Fundamental and applied biological sciences. Psychology Health. Pharmaceutical industry HIV-1 Human immunodeficiency virus 1 Industrial applications and implications. Economical aspects Insect cells Microfiltration Molecular weight Other active biomolecules Production of active biomolecules Purification Ultrafiltration Virus-like particles Viruses |
| title | Characterization and downstream processing of HIV-1 core and virus-like-particles produced in serum free medium |
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