Use of flow cytometry to monitor infection and recombinant human alpha-1,3/4 fucosyltransferase production in baculovirus infected Sf9 cell cultures.
This paper describes the setup and the use of a flow cytometric method for monitoring Sf9 insect cell infection by a recombinant baculovirus expressing the human alpha1,3/4 fucosyltransferase Fuc-TIII. Using side scattered light coupled to green fluorescence detection after immunolabeling of the recombinant protein, this method made it possible to monitor baculovirus infection of Sf9 cells grown in batch cultures and infected at different cell densities and multiplicities of infection. The method was able to precisely assess the extent of infection of the insect cells from 60 h postinfection. In asynchronously infected Sf9 cell cultures, the two-step infection process (primary and secondary infection) was well-characterized using this technique. Finally, a reduced sensitivity to baculovirus infection was observed for cells infected at the end of the growth phase compared to the cells infected during exponential growth phase.
Deparis
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Laboratoire des Sciences du Génie Chimique-CNRS, Institut National Polytechnique de Lorraine-ENSAIA, 2, avenue de la Forêt-de-Haye, BP 172, F-54505 Vandoeuvre-lès-Nancy, France.
Jestin
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A
Marc
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Goergen
J L
JL
D000818:Animals; D016367:Baculoviridae / Q000254:growth & development* / Q000302:isolation & purification / Q000472:pathogenicity*; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D005434:Flow Cytometry / Q000379:methods*; D005647:Fucosyltransferases / Q000032:analysis / Q000096:biosynthesis*; D006801:Humans; D018411:Spodoptera / Q000201:enzymology* / Q000821:virology*
D005647:Fucosyltransferases; C019818:3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase