A review on preparative and semi-preparative offgel electrophoresis for multidimensional protein/peptide assessment

•A review on preparative and semi-preparative offgel electrophoresis.•Offgel electrophoresis trends.•Protein pellet treatments for offgel electrophoresis.•Offgel electrophoresis coupling with electrophoretic/chromatographic techniques.•Offgel electrophoresis for metal–protein complexes assessment.

Mass spectrometry (MS) techniques are commonly used for protein identification and further analysis of selected protein spots after high resolution 2-D electrophoresis. Complementary gel-free approaches have been developed during the last few years and have shown to be useful tools in modern proteomics. The development and application of various gel-free electrophoresis devices for performing protein fractionation according to the pI differences is therefore a topic of interest. This review describes the current state of isoelectric focusing (IEF) gel-free electrophoresis based on the Agilent offgel 3100 fractionator. The review includes, therefore, (i) an overview on IEF as well as other previous IEF gel-free electrophoresis developments; (ii) offgel fundamentals and future trends; (iii) advantages and disadvantages of current offgel procedures; (iv) requirements of isolated protein pellets for further offgel fractionation; (v) offgel fraction requirements to perform the second dimensional analysis by advance electrophoresis and chromatographic techniques; and (vi) effect of the offgel operating conditions on the stability of metal–protein complexes.

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