Figure 1.

BChE/hSA fusion protein expression, activity and oligomeric forms. (A) BChE activity gel staining of samples from BChE/hSA produced in vitro and in vivo. Unless otherwise noted 15 μl (10 units/ml) of samples were loaded onto each lane of a precast 4–20% Tris-Glycine polyacrylamide non-denaturing gel in the following order: lane 1, purified plasma huBChE (tetramer indicated); lane 2, Harvest 8 of conditioned media of clone 35 from the hollow fiber system; lane 3, control conditioned media; lane 4, diluted milk sample from a F1 transgenic mouse, 307-1A7; lane 5, diluted milk sample from a F2 transgenic mouse, 307-1A7A4; lane 6, diluted milk sample from a nontransgenic FVB mouse (1:30); lane 7, diluted milk sample from a transgenic goat, 2237; lane 8, diluted milk sample from a transgenic goat, 2177; lane 9, purified BChE/hSA sample from milk of the goat 2177; lane 10, diluted milk sample from a nontransgenic goat (1:30). (B) Western blot analysis under denaturing and reducing conditions. Immunodetection was performed with a polyclonal anti-huBChE antibody (Dako, Mississauga, ON, Canada). Unless otherwise noted 15 μl (10 units/ml) of samples were loaded onto each lane of a precast 4–20% Tris-Glycine polyacrylamide gel in the following order: lane 1, Biotinylated molecular markers (Cell Signaling Technology, Inc., Danvers, MA); lane 2, purified plasma huBChE; lane 3, Harvest 8 of conditioned media of clone 35 from the hollow fiber system; lane 4, control conditioned media; lane 5, diluted milk sample from a F1 transgenic mouse, 307-1A7; lane 6, diluted milk sample from a nontransgenic FVB mouse (1:30); lane 7, diluted milk sample from a transgenic goat, 2237; lane 8, diluted milk sample from a transgenic goat, 2177; lane 9, purified BChE/hSA sample from milk of the goat 2177; lane 10, diluted milk sample from a nontransgenic goat (1:30). (C) SEC-HPLC analysis of milk from a transgenic mouse, 307-1A7A2. (D) SEC-HPLC analysis of milk from the transgenic goat, 2177.