Cell source Stimulation Secretion stop Fixation Permeabilisation Staining Protocol



That is what it is all about; STAINING. Showing the target is there and with a bit of luck you can pin point the place the target is in (using fluorescence microscopy). The staining is depending on all the other factors; cells, inhibition of secretion, fixation, permeabilization and the antibodies. 
The staining can be performed at room temperature. Since the cells are stabilized by the fixative. And the diffusion is higher with a higher temperature. This way antibodies can go more easily in and out of the cells. Some fluorochromes are temperature sensitive though, like the PerCP conjugates. Should you use this kind of fluorochromes incubation on a bed of ice or in the fridge is the solution. Incubate in all cases in the dark. That way you fluorochromes last longer.

And when you think you thought of everything something went wrong......... We are still working on the 'chapter' of trouble shooting.


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Page was last updated: 05-03-2005