The bound antibodies were detected using the SuperSignal West Dura Trial Kit (Thermo Scientific, Rockford, IL, USA).
Detection was achieved with the appropriate secondary Abs coupled to HRP, followed by ChemiLucent Trial Kit (Chemicon Int).. Immunoblotting with rabbit polyclonal Abs to Akt, IkBα and actin (all dilution 1/1000) (all from Cell Signalling Technology) were used as internal loading controls to ensure equivalent amounts of protein in each lane.
Finally, the blots were detected by enhanced chemiluminescence (ECL) using the LumiGlo substrate (Super Signal West Pico Trial Kit, Pierce, USA).
Antibodies bound to GFP protein, Rubisco large subunit protein and aFGF protein were visualized by chemiluminescent detection (SuperSignal WestPico Trial kit from Pierce).
Then the blot was washed and detected by enhanced chemiluminescence (ECL) using the LumiGlo substrate (Super Signal West Pico Trial Kit, Pierce , USA.
Immunoreactive bands were visualized with Western blotting luminal reagents (Western Blotting Luminol Reagent sc-2048 Trial Kit, Santa Cruz Biotechnology Inc., CA, USA).
Chemiluminescent signals were generated using a SuperSignal West Pico trial kit (Pierce) and were detected by exposure to X-ray film or using the ChemiDoc XRS System (Bio-Rad Laboratories).
The membranes were conjugated with HRP-labeled secondary antibody, and the signals were detected using SuperSignal® West Femto Trial Kit Prod #34094 (Thermo Scientific, Rockford, IL).
After incubation, the protein bands were visualized using a SuperSignal West Pico Trial Kit (Pierce, Rockford, IL) and exposed to radiographic film.
Each strip was mixed with 80 µl of a mix (1 1) of Ultra Supersignal for 5 min. Proteins were visualised by enhanced chemiluminescence (SuperSignal West Dura Trial Kit, Pierce, Rockford, Illinois, USA).
Finally, the blot was washed and detected by enhanced chemiluminescence's (ECL) using the LumiGlo substrate (Super Signal West Pico Trial Kit, Pierce, USA), and the signals were recorded by an imaging System (Bio-Rad, USA), and analyzed with Quantity One software (Bio-Rad, USA).
