When performing 2-DE studies of complex protein mixtures, the nature of the analyzed samples, the pI range of their proteins as well as the purpose of the study have to be considered. With regard to the aim of generating a proteome analysis of human saliva and of establishing a high-resolution bacterial overlay it turned out that focusing over a pH range of 3-10, by in-gel rehydration, is the best way to display the whole range of salivary proteins in a reproducible way. In terms of sample preparation, a compromise has to be found between a high number of protein spots and the true display of all typical salivary proteins, thus filtrating of WS prior to electrophoresis experiment was chosen as the method of choice for subsequent studies.
Acknowledgements
I’m especially grateful to Verena Wittmann, who was my first instructor for two-dimensional gel electrophoresis as well as Bernhard Gess, Institute of Physiology, University of
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Chapter 4 Proteome Analysis of Human Saliva