For extraction of proteins I freeze tissues on liquid Nitrogen and grind them in a mortar with a pestle. I then add about 10 mls of ice cold homogenizing buffer per 1 gram of tissue. I have found that grinding plant tissues with a mortar and pestle works better than nitrogen cavitation or a dounce homogenizer due to the difficulty in breaking up the cell wall.
Depending on what protein you are looking at, you may want to try a couple of different time points. For example you may want to try tissue samples from cultures 3-4 days after transfer if your protein is growth associated or 7 days after transfer if your protein expression is associated with a quiescent state. If you want a more detailed protocol, send E-mail to me. By the way, do you know how I can attain Tobacco BY-2 cells. We have not used the cell line for quite some time now and I would like to do a few more experiments. Unfortunately, our cell line as ceased to exist.
Ken MaasDept. of Biological SciencesNorthern Illinois UniversityDeKalb, Illinois USA email@example.com