Purification and analysis of infectious virions and native non-structural antigens from cells infected with tick-borne encephalitis virus

The NS1 protein of tick-borne encephalitis virus forms multimeric species upon secretion from the host cell

“…Previous observations from the purification of extracellular antigens (Crooks et al, 1990) has suggested that NS1 may exist in forms with larger M r values than the monomers or dimers normally found in infected cells and their supernatants. Research on the structure of NS1 has so far depended on immune precipitation followed by PAGE or on analysis by Western blotting.…”

“…Infected cell supernatants were clarifed, concentrated and analysed on Sephacryl $300 (Pharmacia) columns in PBS pH 8.0 as described previously (Crooks et al, 1990). The columns were calibrated with high M r markers (Pharmacia) according to the manufacturer's instructions.…”

“…Although little is known about the structure and function of this protein, it has been shown to exist as a dimer in cells infected with every tick-borne or mosquitoborne flavivirus so far studied (Winkler et al, 1988;Jacobs et al, 1992;Flamand et al, 1992), although preliminary data from this laboratory suggest that the extracellular form may be even larger (Crooks et al, 1990). In addition, its primary structure is highly conserved (Jacobs et al, 1993;Venugopal et al, 1994), especially the N and C termini, although the central region is relatively variable.…”

Chromatographic purification of recombinant adenoviral and adeno-associated viral vectors: methods and implications

“…Size-exclusion chromatography, also called gel filtration, is the method of choice for purification of many viruses, including tick-borne encephalitis virus and retrovirus. 64,65 It can also be used as a polishing step for Ad purification. 45 A few disadvantages are associated with the use of size-exclusion chromatography for virus purification.…”

Maturation of the dengue-2 virus NS1 protein in insect cells: effects of downstream NS2A sequences on baculovirus-expressed gene constructs

“…Present address: Institut Pasteur, Laboratoire des Arbovirus, 25 rue du Dr Roux, 75724 Paris Cedex 15, France. Fax + 33 1 45688780. e-mail cprehaud@mendel.sis.pasteur.fr (Winkler et al, 1988), although a hexameric form has been described by Crooks et al (1990). Newly synthesized monomeric NS1 is a hydrophilic protein which becomes partially hydrophobic and membraneassociated during or shortly after dimerization (Winkler et al, 1989).…”