title: [Expression of murine interleukin 12(mIL-12) in insect cells].
abstract: Since human IL-12 is species-specific in its functions and elicits little biological responses from mouse lymphocytes, it is necessary to express recombinant murine IL-12 for the usage in studying the effects of this cytokine in various rodent models. Thereby, we can investigate the role of IL-12 in immune response in vivo and evaluate its potential clinical utility. Thus, we firstly constructed two expression vectors, pVL1393-mp40 and pVL1393-mp35. They were used to co-transfect the insect cells(Sf9) separately with linearized polyhedrosis virus genomic DNA. Two kinds of recombinant viruses AcNPV-mp40 and AcNPV-mp35 were visually screened out, and mp40 and mp35 were co-expressed in the insect cells co-infected by AcNPV-mp40 and AcNPV-mp35. The results of real-time Biomolecular Interaction Analysis (BIA) and Northern blot demonstrated that the recombinant mIL-12 was expressed successfully in the insect cells. The molecular weights of recombinant mp40 and mp35 were 40 KDa and 22 KDa on SDS-PAGE under reducing conditions, respectively. The apparent molecular weight of recombinant mIL-12 is 80 KDa under non-reducing conditions of Western blot. Biological activity of the recombinant product was detected in conditional medium using antibody-capture bioassay. The expression level of recombinant mIL-12 was about 10-15 micrograms/10(6) cells, as compared with the calibration curve of mIL-12.
authors.affiliation: Shanghai Institute of Cell Biology, Academia Sinica, Shanghai 200031.
authors.forename: C L
authors.initials: CL
authors.lastname: Li
authors.forename: H
authors.initials: H
authors.lastname: Shen
authors.forename: M Q
authors.initials: MQ
authors.lastname: Lin
authors.forename: Z L
authors.initials: ZL
authors.lastname: Zhang
authors.forename: Q X
authors.initials: QX
authors.lastname: Shen
mesh_terms: D000818:Animals; D018664:Interleukin-12 / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000096:biosynthesis; D018411:Spodoptera / Q000166:cytology / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D018664:Interleukin-12