This PhD thesis is focused on the molecular weight determination and systematic characterization of proteins and poly(amido amine) (PAMAM) dendrimers with molecular weights higher than 200 kDa using two gel electrophoretical (planar SDS-PAGE and CGE-on-the-chip) and one mass spectrometric technique (MALDI-TOF-MS). For this purpose a novel high molecular weight protein assay for CGE-on-the-chip was developed.
CGE-on-the-chip is the most recent of all three applied analytical techniques and was so far only applied for the analysis of proteins with molecular weights lower than 250 kDa. Analysis of high molecular weight proteins with MALDI-TOF-MS was only sparely described in literature. In this work it was demonstrated that all three described analytical techniques were capable of analyzing high molecular weight protein samples. The results obtained with CGE-on-the-chip were in very good agreement with the protein bands found with traditional planar SDS-PAGE.
For all high molecular weight proteins exact molecular weights could be determined with MALDI-TOF-MS using a commercial standard instrument.
Another issue was the exact molecular weight determination of PAMAM dendrimers using MALDI-TOF-MS. So far only the molecular weight of smaller dendrimers was determined with MALDI-TOF-MS. It was shown that exact molecular mass determination of PAMAM dendrimers up to generation 10 was generally possible using MALDI-TOF-MS. Moreover it was found out that the polydispersity of the analysed dendrimer samples was significantly higher than the polydispersity of proteins in the same mass and size range. Due to imperfect dendrimer growth/synthesis the molecular weights determined with MALDI-TOF-MS for PAMAM dendrimers were significantly lower than the expected molecular weights.