To evaluate the application of reverse staining in proteomic research, the same protein samples were run by two-dimensional electrophoresis, then the gels were dyed by reverse staining and coomassie Brilliant Blue (CBB) staining. The reverse staining and CBB staining gels were compared, eight pairs corresponding protein spots from the two different gels were cut out, digested by trypsin and analyzed by MALDI-TOF/ TOF. Image analysis showed that the reverse staining gel could display more protein spots than the CBB staining one. Seven of eight protein spots on reverse staining gel were effectively identified by MALDI-TOF/TOF, and eight protein spots on CBB staining gel. So, we consider that the sensitivity of reverse staining is higher than the CBB staining, and its compatibility with MALDI-TOF/TOF is good. Therefore, the reverse staining method can be used to set up the reference map of 2-DE, but not suitable to use on the comparative of proteins.
