Protamine is a poly arginine peptide, It is used for transferring DNA into cells (together with liposomes most of the times). Cells take up the protamine by endocytosis. In the literature they incubate the cells with protamine for 6 hours or more. Is incubating the cells for 15 min with protamine enough for endocytosis?
The protamine will likely be taken up by a mixture of different endocytic mechanisms including clathrin-mediated, non-clathrin-mediated and processes such as macropinocytosis - although the use of liposomes might promote a non-endocytic uptake mechanism involving membrane fusion. How quickly it is taken up will depend on how actively the various uptake mechanisms are operating in the cells you are intending to transfect. My guess is that some cells will transfect in 15 mins or even less but you will have more efficient transfection(s) is you leave the protamine on the cells for longer.
I'm sorry I can't be more precise but the question can't really be answered much more precisely with the information you have provided. This is one example where empirical science may give you an answer, i.e. try different incubation times with your cells and see which gives you the best results.
Sorry I forgot to determine the cells. These are V79 (Chinese hamster fibro blast cells) and CHO (Chinese hamster ovary cells).
Rate of endocytosis is highly dependent upon the factors like, incubation conditions which include; temperature, size of cargo (in your case liposome), charge of cargo, cell type etc.
Liposomes endocytosed the same way like any other cargo and this is truly a misconception that they fuse with lipidbilayer. I never worked with CHO but if I were you, I will search literature about optimum measures I should take with these cells for quicker endocytosis.
From my experience, I can tell that generally half an hour is enough at 37degree for seeing any signal inside the cells. increasing time to 3 h will increase the signal as more material (if not removed from medium) will be endocytosed. you are working with DNA transfection, taking time in account does make sence in such type of experiment. you can run an experiment with 15 min, 1/2 h, 3 h and see which time is suitable for your particularly need. I never tried 15 min and dont know anyone else from litrature who did transfection for 15 min using only endocytosis.
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