Production of Influenza Virus-like Particles from Stably Transfected Trichoplusia ni BT1 TN-5B1-4 Cells

초록

We investigated the production of influenza A/Korea/01-2-9/2009 (H1N1) virus-like particles (VLPs) containing four structural proteins, matrix protein 1 (M1), matrix protein 2 (M2), neuraminidase (NA) and hemagglutinin (HA), using stably transfected Trichoplusia ni BT1 TN-5B1-4 (TN-5B1-4) cells. Recombinant M1, M2, NA and HA were expressed as bands with molecular weights of 28, 17, 60, and 70 kDa, respectively, in stably transfected TN-5B1-4 (TN-5B1-4/M1-M2-NA-HA) cells. VLPs were purified from the culture medium of the TN-5B1-4/M1-M2-NA-HA cells by pelleting on a sucrose cushion, followed by ultracentrifugation in a sucrose density gradient. The four structural proteins were released together from the TN-5B1-4/M1-M2-NA-HA cells, and were co-purified from the same fractions of the sucrose density gradient, indicating that recombinant M1, M2, NA, and HA self-assembled into VLPs in the TN-5B1-4/M1-M2-NA-HA cells. Recombinant baculoviruses (rBac/NA and rBac/HA) expressing recombinant NA and HA were generated and used to co-infect stably transfected TN-5B1-4 (TN-5B1-4/M1-M2) cells expressing recombinant M1 and M2, resulting in the production of high-molecular-weight VLPs and the release of self-assembled VLPs with diameters of 80 similar to 120 nm. The production of VLPs was greatly enhanced in the TN-5B1-4/M1-M2 cells co-infected with rBac/NA and rBac/HA, compared with the TN-5B1-4/M1-M2-NA-HA cells. Our results suggest that a stably transfected-insect cell expression system might be useful for producing VLPs for the development of recombinant vaccines against influenza.

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