title: Ion-exchange membrane chromatography method for rapid and efficient purification of recombinant baculovirus and baculovirus gp64 protein.
abstract: Significant progress in the application of baculoviral vectors for gene delivery into mammalian cells calls for the development of powerful methods for virus purification and concentration. We report here a novel and efficient method based on membrane chromatography to prepare baculoviral stocks. On a strong cation-exchange membrane unit, baculovirus in insect cell culture supernatants was captured at a flow rate of 3 ml/min and efficiently eluted at the same flow rate with a physiological buffer containing 150 mM NaCl as a desorption reagent. The procedure allowed for a final recovery of 78% of infective viral particles from the original supernatant and 30-fold enrichment. The high purity of the viral preparation was demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. Baculovirus gp64 proteins could be purified by the same method, indicating the importance of the protein in mediating the binding of baculovirus to the cation-exchange membrane. The method developed should be suitable for preparing baculoviral stocks, and probably other gp64-pseudotyped viral vectors, for gene therapy applications.
authors.affiliation: Institute of Bioengineering and Nanotechnology, Singapore.
authors.forename: Chunxiao
authors.initials: C
authors.lastname: Wu
authors.forename: Ker Yin
authors.initials: KY
authors.lastname: Soh
authors.forename: Shu
authors.initials: S
authors.lastname: Wang
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D002460:Cell Line; D002852:Chromatography, Ion Exchange / Q000379:methods*; D005822:Genetic Vectors / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D018411:Spodoptera / Q000821:virology; D013997:Time Factors; D014763:Viral Matrix Proteins / Q000302:isolation & purification*; D015678:Viral Structural Proteins / Q000302:isolation & purification*
chemical_list: D011994:Recombinant Proteins; D014763:Viral Matrix Proteins; D015678:Viral Structural Proteins; C065417:major structural protein, cytomegalovirus 64kDa