Investice do rozvoje vzdělávání Overview of the use of MALDI-TOF MS and MALDI MSI within the NANOTEAM project Roman Guráň 17.7.2015 Investice do rozvoje vzdělávání

Bruker New ultrafleXtremeTM The innovative smartbeam-II™ laser enables ultra-high data acquisition speed in both MS and MS/MS at full systems performance. The well-established proprietary smartbeam laser provides unprecedented analytical and matrix flexibility in workflows from protein tissue imaging, intact proteins analysis, Glycoproteomics, biologics or oligo QC, to LC-MALDI proteomics - all fully enabled at 1-2000 Hz repetition rates. The new ultrafleXtreme now enables laser focus diameters down to 10 µm for high spatial resolution imaging without pixel overlap. Importantly, outstanding spectral quality and signal intensity are maintained at even the smallest laser beam diameters. The novel FlashDetector™ combined with a minimum of 4 GHz digitizer and latest advances in electronics provide unmatched mass resolving power up to 40,000 and 1 ppm mass accuracy for highest confidence. The novel and unique laser-irradiation self-cleaning Ion Source ensures robust, long-term highest-performance operation. Very long MALDI laser lifetime in combination with automated source cleaning in just minutes leads to high uptime and low maintenance costs¨. Latest TOF/TOF technology: The high efficiency and sensitivity of the LID-LIFT process delivers MS/MS spectra with nominal mass resolution for peptides. Typically, full MS/MS data sets can be acquired with up to 1000 Hz laser repetition rate from low fmol levels within seconds. The Bruker bioinformatics software is perfectly adapted to analyze and visualize the match between the raw spectra and annotated peptide and protein structures.

Applications in our lab Identification of proteins, bacteria and other MALDI mass spectrometry imaging of different tissues

Results under CEITEC project Influence of microbiome species representation in the hard-to-heal wounds on disease severity and treatment duration MALDI-TOF MS was used for identification of bacteria and then the dendrograms were created for comparing identified bacteria.

Results under CEITEC project Structural effects and nanoparticle size are essential for quantum dots-metallothionein complex formation MALDI-TOF MS was used for characterization of metallothionein. Peak No. Molecular mass found (Da) Molecular mass expected (Da) Identification 1 6111 6113 Apo MT-2C 2 6131 - 3 6139 N-acetylated and C-methylated apo MT-2A 4 6171 6169 N-acetylated and C-methylated apo MT-2C 5 6210 6209 Na+ ion of C-methylated apo MT-2D 6 6231 6230 N-acetylated and C-methylated apo MT-2D 7 6321 6322 adduct of Na+ ion of N-acetylated apo MT-2B with molecule DHB (matrix) 8 6362 6363 adduct of Na+ ion of C-methylated apo MT-2D with molecule DHB (matrix) 9 6418   6417 adduct of Na+ ion of N-acetylated apo MT-2E with molecule DHB (matrix) adduct of N-acetylated and C-methylated apo MT-2D with molecule DHB (matrix) B Intensity (a.u.) 6 000 6 200 6 400 6 600 2 000 4 000 8 000 5 800 1 5 4 6 7 23 8 9 m/z

Results under CEITEC project Differences in urinary proteins related to surgical margin status after radical prostatectomy MALDI-TOF MS was used for identification of urinary proteins by peptide mass fingerprinting (PMF) after 2D gel electrophoresis.

Results under SIX project Study of Linkage between Glutathione Pathway and the Antibiotic Resistance of Escherichia coli from patients’ swabs MALDI-TOF MS was used for comparison of different E.coli samples and for detection of glutathione (GSH).

Results under SIX project Growth properties and ability to form biofilms by Staphylococcus aureus and MRSA after the treatment of antibiotics with SeNPs MALDI-TOF MS was used for comparing protein profiles of S. aureus and MRSA after application of antibiotics with SeNPs.

Results under SIX project The substitution of organic matrices by carbon nanomaterials for MALDI-TOF MS analyses of low molecular weight analytes like glutathione MALDI-TOF MS was used for testing carbon nanomaterials as new matrices for low molecular weight analytes.

Future work MALDI MSI of different tumors – we will focus especially on detection of metallothioneins Identification of other bacterial samples from Trauma hospital of Brno

Investice do rozvoje vzdělávání Acknowledgements Investice do rozvoje vzdělávání 11

Thank you for your attention Investice do rozvoje vzdělávání