Automation of In-gel digestion and MALDI spotting of two-dimensional electrophoresis separated rat liver proteins
2005
Tenz,K. | Steenvorden,R. | Krause,E. | Paal,K. | Oberemm,A.
Alemán. Proteomic approaches are widely explored in short term toxicological tests and for gaining insight into toxic and carcinogenic effects of chemicals. Since emerging proteomic technologies still depend on data generated by two-dimensional electrophoresis (2-DE), our aim was to establish a new 2-DE based map of male Wistar rat liver proteins in the context of a molecular toxicological joint research project. At present, the map contains 621 proteins and could be the basis for a publicly accessible HTML-database, providing quick information to identify protein expression patterns of toxicological relevance. As an alternative to our approved manual procedure (optimized at the FMP) high performance sample preparation requires a robot system capable of spot picking, digesting and spotting directly onto MALDI target. The best results has been delivered by the ProPic spotpicker in combination with the ProGest digester and ProMS spotting station (all from Genomic Solutions) allowing simultaneous processing of 96 gelspots within 12 hours. The ProMS is compatible with the most commercially available MALDI targets. We have successfully transferred our manual protocol to the automation system and tested strongly as well as weakly stained spots and both led to confidential spectra with high peak intensities and identification scores.
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