Threshold of 0.02. The analysis Adriamycin Doxorubicin of 1D annotation is more informative, if on a big number of entries s F tions per semester Calculated llig annotation in the design of the statistical analysis. Therefore, the proteins were With at least one event of quantification for analysis annotation as D. histograms for the median normalized ratio Ratios of proteins between 17 DMAG treated and controlled The statistical were plotted using the R programming language. , In the calculation of the correlation coefficient data records Tze for valid entries GE with quantification in all experiments filtered. Proteins which are the t-test have been passed in Table 2, further, the ratios, the corresponding ratio Summarized in each repetition. RESULTS quantify the coverage of the proteome in depth to them Changes by 17 DMAG in HeLa cells induced proteome, we used SILAC-based quantification and high resolution and high MS coupled to LC / MS. A suitable concentration of 17 DMAG was found at 50 and proteomics Ver Changes 24 hours, after drug treatment, when Analyzed changes in protein levels at the st Appeared to be strongest. We analyzed five biological replicates, where equal amounts of protein from the control sample DMAG and 17-treated HeLa cell lysates were mixed, digested with trypsin in L Solution supports using the filter by methods of preparing samples and the resulting peptide by strong anion exchange chromatography-based pipette. The six peptide fractions repetition of each experiment were then analyzed by liquid chromatography-mass spectrometry measurement in tandem, a hybrid LTQ Orbitrap high Aufl With more energy Sto Dissociation fragmentation of the base. The combined data set more than five repetitions proteome includes 30 LC MS / MS with a slope of 4 h in contrast to the conventional strategy of low-rise, where aremeasured obtain fragments in a linear ion trap, we have a level of two ppm for precursor Shore- ions and their fragments completely through the acquisition of two requests reference requests getting scans and MS MS / MS scans in the Orbitrap analyzer fragmentation. Examples of the Datenqualit t is illustrated in Figure 1B, the HCD-based MS / MS spectrum of doubly charged peptides of heat shock protein Hsp27 and EGFR receptor tyrosine kinase, which is characteristic of the extensive coverage of sequence ions y and some B-ions of low mass.
The high mass accuracy of fragment ions is shown by the low ppm error for measurements Orbitrap. Analyzed more than 60% MS / MS and reliably Permeable, which then causes no identification 7000 proteins Tested at a rate of false detection of proteins of less than 1%. Protein identification was highly reproducible among replicates with more than 85% of the proteins, the judge in at least three repeated experiments, we identified the depth of proteome to the increased impact of the 17 Ma DMAG. Quantification of supply changes Of proteomic data allows a pr Precise quantitative comparison of approximately 6000 proteins in response to inhibition axitinib of Hsp90 at 17 DMAG treatment. A summary of the identification and quantification of proteins in individual experiments is shown in Table 1 erg Complementary. SILAC quantification of peptides is measured, with peptides corresponding to EGFR and Hsp27, both of which are known to be influenced by the inhibition of Hsp90 by 17 DMAG shown. W While EGFR.