when spectra from all sample subtypes are included directly in CPT and standard analysis is performed. It is noteworthy that when using DHB as a MALDI matrix NBI-56418 provided a higher number of mass peaks as compared to CHCA. Likewise, the Ga-based IMAC approach produces more mass signals as compared to the Fe-based assay. In addition, the peak lists derived from DHB spectra showed a higher mean correlation between data sets. These results suggest that MALDI analyses using Ga-based IMAC and DHB as MALDI matrix are more reproducible and provide a higher number of mass signals. The peaks identified derived from highly expressed proteins and the remaining discriminating peptides could not be identified by MALDI MS. Alternative identification strategies should be tested in order to increase identification of low-intensity signals in MALDI MS studies. Discriminant analyses allowed us to separate normal lung and NSCLC samples and to identify the peptides which best 1161233-85-7 structure discriminated between normal and diseased tissues, as shown by clustering analysis. However, this task is not usually problematic due to the important differences between normal and cancer tissues. What proves trickier is finding differences between distinct histological subtypes. As showed in Figure 1, there are two main clusters of lung cancer samples, including adenocarcinomas and large cell carcinomas separately, but squamous cell carcinoma samples are splitted between these clusters. It has been described that ensemble classifiers outperform single decision trees classifier by having greater accuracies and smaller prediction errors when applied to proteomics datasets. So, we tested if AdaBoost analyses could classify the different NSCLC samples correctly. Our results suggest that AdaBoost can discriminate samples of one lung cancer histological subtype from the other two. The use of technical replicates as test set allowed us to assess the robustness of the methodology employed. Our data suggest that both GAPDH and b-actin have a significantly increased expression in lung cancer samples. Overexpression of GAPDH in human lung cancers was described previously by Tokunaga et al and there are many publications showing increased expression of GAPDH in breast, pancreatic and cervical human cancers. On the other hand, several studies indicated that b-actin was differentially expressed in human cancer. Both proteins showed increased levels in rat hepatoma. Moreover, IHC expression profiles for b-actin and GAPD
