Successful protein spot identification by MALDI-TOF MS in conjunction with PMF and/or Q-TOF MS/MS with BLAST database searching.

1Statistically significant protein spot identification after MALDI-TOF MS analysis followed by PMF search of the H. contortus putative EST protein database.
2Non-statistically significant protein spot identification after MALDI-TOF MS analysis followed by PMF search of the H. contortus putative EST protein database, but verified by statistically significant protein spot identification after Q-TOF MS/MS analysis.
3Statistically significant protein spot identification after MALDI-TOF MS analysis followed by PMF search of the H. contortus putative EST protein database. These results were confirmed by Q-TOF MS/MS analysis.
4Statistically significant protein spot identification after Q-TOF MS/MS followed by BLASTp search of the H. contortus putative EST protein database.
5Non-statistically significant protein spot identification after MALDI-TOF MS analysis followed by PMF search of the H. contortus putative EST protein database. However, the observed Mw of the spots, calculated on the gel image, were correlated with the theoretical Mw of the intact protein in the best BLASTp match. These data are detailed in Tables S3 & S4.
Each protein spot was excised from the 250 µg protein-loaded gel and analysed by MALDI-TOF MS. A local MASCOT PMF search of the H. contortus putative EST protein database was performed and the highest scoring EST sequence match, along with its MOWSE-based score (significance threshold score >51, p-value<0.05) is reported. 20 spots were also selected for Q-TOF MS/MS, excised from the 500 µg protein-loaded gel and analysed by Q-TOF MS/MS. A local MASCOT Ion Search of the H. contortus putative EST protein database was performed and the MOWSE-based score of the highest scoring EST sequence match (significance threshold score >25, p-value<0.05) are given in brackets. Following either method, each EST sequence hit was submitted to a BLASTp search against the entire NCBI nr protein database. For each search, the highest scoring hit score (significance threshold >44, p-value<0.01), its accession number and protein name are reported. For information, the species corresponding to the highest scoring hit and the method of identification used are also given.