The freeze-dried samples were diluted with sterile distilled water in order to obtain 1 μg of total protein/μL. To preserve proteins from enzymatic degradation, the dilutions were immediately stored at -20°C until use. Five μg of sample were first diluted (1/20) in binding buffer and loaded on CM10, Q10, H50 and IMAC30-Cu2 or IMAC30-Zn2 ProteinChip then incubated for 1hr at room selleck temperature. The unbound proteins were removed by washing three times with 200 μL of the same buffer, the ProteinChips® were quickly rinsed with pure water and left to dry. For NP20 ProteinChips® , 2 μL of sample were applied

on the spot and left to dry, and then washed three times with 5 μL of water. Matrix (100% saturated solution of sinapinic acid in 0.5% trifluoroacetic acid/50% acetonitrile) was applied to each spot (twice 0.8 μL). The absorbed proteins were then analyzed on a ProteinChip Reader (series 4000, Bio-Rad Laboratories, Hercules, CA, USA). Spectra were obtained using two different acquisition protocols, for low (2.5-14 kDa) and high (14-400 kDa) molecular mass proteins, respectively. External mass calibration was performed with ProteinChip All-in-One SB203580 price Protein

Standard II (Bio-Rad, laboratories, Hercules, CA, USA). Peak annotation was performed after base-line subtraction, noise calculation, and normalization by total ion current (TIC). Peak detection was achieved with ProteinChip Data Manager Software and only peaks with a signal-to-noise ratio > 5 were used for analysis (Bio-Rad Laboratories, Hercules, CA, USA). Statistical analysis Statistical analyses were performed using ProteinChip Data Manager 3.0 software (Bio-Rad Laboratories, Hercules, CA, USA). All the spectra were compiled, and qualified mass peaks (signal-to-noise ratio > 5)

with mass-to-charge ratio (m/z) between 2.5 kDa and 250 kDa were auto detected. P-values were calculated using non parametric Mann-Whitney U-test, which tests the null hypothesis that the medians of the peak intensities of the groups are equal. A p-value less than 0.05 was accepted as statistically significant. The difference was also examined by hierarchical clustering. Acknowledgements and funding Clomifene We gratefully thank Christel Binard and Sabine Durville for reading the manuscript and improving the English redaction. This study was supported by the Belgian Science Policy Office (contract C3/00/19). References 1. Latgé JP: Aspergillus fumigatus and aspergillosis. Clin Microbiol Rev 1999, 12:310–350.PubMed 2. Latgé JP: The pathobiology of Aspergillus fumigatus . Trends Microbiol 2001, 9:382–389.PubMedCrossRef 3. Geiser DM, Klich MA, Frisvad JC, Peterson SW, Varga J, Samson RA: The current status of species recognition and identification of Aspergillus . Stud Mycol 2007, 59:1–10.PubMedCrossRef 4. Hohl TB, Feldmesser M: Aspergillus fumigatus : principles of pathogenesis and host defense. Eukaryotic Cell 2007, 6:1953–1963.PubMedCrossRef 5.