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Establishment of a Large-Scale Purification Procedure for Purified Recombinant Bovine Interferon-.TAU. Produced by a Silkworm-Baculovirus Gene Expression System

2004; Japanese Society of Veterinary Science; Volume: 66; Issue: 11 Linguagem: Inglês

10.1292/jvms.66.1395

1347-7439

Hidekazu Nagaya, Toshimichi Kanaya, Hiroki Kaki, Yoneko Tobita, Masashi Takahashi, Hitomi Takahashi, Yuichi Yokomizo, Shigeki Inumaru,

Virus-based gene therapy research

We developed a procedure for the large-scale purification of bovine interferon-τ (boIFN-τ) by means of a silkworm-baculovirus gene expression system. Recombinant boIFN-τ (rboIFN-τ) was efficiently produced in the silkworm infected with boIFN-τ cDNA recombinant baculovirus and accumulated in the haemolymph. To establish a purification method suitable for mass production, we tried three crude purification methods, namely, an acidification and neutralization treatment (ANT), silica gel column chromatography (SGCC), and Blue sepharose column chromatography (BSCC) with a combination of Q-sepharose (QSC) and chelating sepharose column chromatographies (CSCC). As a result, the acidification and neutralization treatment was found to be the most efficient and cost effective. With this combination, we obtained 91% pure products. To confirm the applicability of the procedure for mass production, we inoculated 100 silkworms with the recombinant virus, and recovered about 4.55 mg (1.26 × 108U/mg) of 91% pure rboIFN-τ by means of a combination of the ANT, followed by QSC and CSCC.