Current Status for High Titre Poxvirus Stock Preparation in CEF Under Serum-Free Medium Conditions: Implication for Vaccine Development

Abstract

In light of the recent detection of BSE in North America and its endemic nature in other regions of the world, there is a real need to employ cell culture conditions that do not require any animal-derived material. Here we report the use of an ultra-low protein serum-free medium (VP-SFM, Invitrogen) for the amplification of poxviruses in primary chicken embryo fibroblasts (CEF). We compared the amplification of four different poxviruses (canarypox, modified Ankara Virus (MVA), vaccinia virus strain Copenhagen and myxoma strain Lausanne) in three different media: DMEM 10%, DMEM 2% and serum-free medium VP-SFM. VP-SFM is a serum-free, ultra-low protein medium containing no proteins or peptides of human or animal origin designed to support the replication of viruses and the production of recombinant proteins and monoclonal antibodies. Our results show that high titre poxvirus stocks can be prepared in VP-SFM equivalent to that prepared in serum containing medium.

An Antitumor Cellular Vaccine Based on a Mini-Membrane IgE

“…Fresh CEF could be passaged three to four times, frozen CEF only one to two times. Serum-free CEF yielded MVA titers of ∼10 7 PFU/ml after 48 h of culture, very similar to the titers produced by serum-supplemented cultures, as already reported (17). Serum-free CEF were used for recombinant virus construction, terminal dilution cloning, and virus stock production, whereas for analytical experiments, DMEM supplemented with 5% FCS, 1% chicken serum (Life Technologies), and antibiotics was used.…”

Cross-protective immunity against multiple influenza virus subtypes by a novel modified vaccinia Ankara (MVA) vectored vaccine in mice

“…Stocks of chicken embryo fibroblasts (CEF) were produced as previously described [20, 21]. The CEF were used for propagating modified vaccinia Ankara (MVA) virus obtained from the Centers for Disease Control, Atlanta, GA.…”

Introduction to animal cell culture technology—past, present and future

“…Whereas the use of serum-free media is largely established for the production of recombinant proteins, the classical viral vaccine production processes still make use of serumcontaining media at least during some phases of the process (Merten 2002) although such processes can be efficiently performed under serum or protein-free conditions, such as for the production of polio virus by Vero cells grown under serum-free conditions (Merten et al 1997). New developments in the vaccine field clearly make use of serum-free media (Merten et al 1996;Brands et al 1999;Kistner et al 1999;Makoschey et al 2002;Gilbert et al 2005).…”