Efficient analysis of protein expression by using two-dimensional electrophoresis (2-DE) data relies on the use of automated image processing techniques. The overall success of this research depends critically on the accuracy and the reliability of the analysis software. In addition, the software has a profound effect on the interpretation of the results obtained, and the amount of user intervention demanded during the analysis. The choice of analysis software that best meets specific needs is therefore of interest to the research laboratory. In this paper we compare two advanced analysis software packages, PDQuest and Progenesis. Their evaluation is based on quantitative tests at three different levels of standard 2-DE analysis: spot detection, gel matching and spot quantitation. As test materials we use three gel sets previously used in a similar comparison of Z3 and Melanie, and three sets of gels from our own research. It was observed that the quality of the test gels critically influences the spot detection and gel matching results. Both packages were sensitive to the parameter or filter settings with respect to the tendency of finding true positive and false positive spots. Quantitation results were very accurate for both analysis software packages.
We have established the first public database of human primary T helper cell proteome using two-dimensional electrophoresis (2-DE) and matrix assisted laser desorption/ionization-time of flight-mass spectrometry. For the database, CD4+ human T cells were activated with anti-CD3+anti-CD28 antibodies and metabolically labeled with [35S]methionine for 24 h. Cells were lysed and proteins were separated by 2-DE. About 1500 protein spots were detected in the resulting 2-DE gels with silver staining, and 2000 spots with autoradiography. We have identified 91 proteins from the 2-DE gels using peptide mass fingerprinting, and annotated them to our database. The identified proteins are also linked to SWISS-PROTand NCBI protein databases. Our database is available via the Internet at http://www3.btk.utu.fi:8080/Genomics/Proteomics/Database.
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