stable transfection - (Feb/17/2006 )
I have done the stable transfection and the expression of my protein is quite low after a few passage. can I do the second times transfection on my stable clone to increase the expression level of my protein? Does anyone did this before?
I don't know if doing another transfection will increase protein expression. In past when I make stable transfectants, I have to grow up many clones and test each clone to see which ones had best characteristics I was looking for. Maybe you are better off starting over with some new cells and plating a lot of cells.
vsoy is right. better to do clones and test which one express the protein as appropriate level. You can do clone establishment and antibiotic selection in one way. You may consider the fact that you won't be able to counter select the twice transformed clones...
Is flow cytometry a possibility, to positively select the higher expressing cells in your population? Depending on the protein you are expressing or if you have it tagged. Just a thought.