Budování výzkumných týmů a rozvoj CZ.1.07/2.3/.00/09.0224 NANOTEAM Budování výzkumných týmů a rozvoj univerzitního vzdělávání výzkumných odborníků pro mikro- a nanotechnologie Seminář na téma: Laboratory of biomarkers and flow techniques Ondřej Zítka 15.4. 2016 Ústav chemie a biochemie, místnost D06

Members of laboratory RNDr. Ondřej Zítka, Ph.D. Mgr. Natálie Cernei, Ph.D. Ing. Jiří Kudr Mgr. Roman guráň Mgr. Markéta Komínková Bc. Zuzana Lacková Bc. Michal Pláteník (technology part) Jan Zítka (technology part)

Activities of laboratory Servis analysis Total aminoacids determination Thiol compounds determination GSH/GSSG, Phytochelatins Isolation of protein using FPLC MALDI identification of proteins MALDI identification of bacteria species Research activities: Developement of analytical methods for study of low molecular metabolites MALDI imaging methods Design and building of Flow injection analysis methods and instruments for isolation and detection of biomolecules using PMPs Design and manufacturing of technology devices

Major research activities Low molecular metabolites– Antioxidants, thiolic peptides, Fytochelatin synthase EC 2.3.2.15 HPLC with electrochemical detection Zitka, O., Krystofova, O., Sobrova, P., et al, (2011c) Phytochelatin synthase activity as a marker of metal pollution. J. Hazard. Mater., 192, 794-800.

Major research activities Low molecular metabolites– Aminoacids, Sarcosine, (Glutamine) Currently analysis for LPR project Ion exchange liquid chromatography

Major research activities Low molecular metabolites– SAM, SAH - potential markers in cancer diagnostics HPLC-MS (Q-TOF) Chromatogram 2 4 6 8 10 Time [min] 1 x10 Intens. SAH,SAM kalibrace_0.004_2.d: EIC 399.4500 +All MS SAH,SAM kalibrace_0.004_2.d: EIC 385.4500 +All MS SAH,SAM kalibrace_0.0004_2.d: EIC 399.4500 +All MS SAH,SAM kalibrace_0.0004_2.d: EIC 385.4500 +All MS SAH,SAM kalibrace_0.00004_3.d: EIC 399.4500 +All MS SAH,SAM kalibrace_0.00004_3.d: EIC 385.4500 +All MS Spectrum View 250.0662 385.1105 SAH,SAM kalibrace_0.004_2.d: +MS, 4.6min #278 100 200 300 400 500 600 700 m/z 298.0844 399.1245 SAH,SAM kalibrace_0.004_2.d: +MS, 2.8min #167 5 800 900 LOD 0,06 µM LOQ 0,21 LOD 0,14 µM LOQ 0,48

Major research activities Proteins – MALDI imaging – under the method developement MALDI – TOF/TOF

Major technology /research activities Low molecular metabolites – Flow injection analysis with paramagnetic particles Building and construction of new fluidic devices Syringe Pump waste Switching valve Detector (CHI Cell) (control) Sample/rinsing buffer inlet PC Dynabeads Protein G Goat Imunoglobulins Mouse Imunoglobulins 8-hydroxyguanosin Step 1: Imunocomplex G Step 2: Imunocomplex G-M Step 3: Imunocomplex G-M-R + Rabbit Anti-mouseImunoglobulins Modified by Alcaline phosphatase Zitka, O., Krizkova, S., Krejcova, L., et al (2011b) Microfluidic tool based on the antibody-modified paramagnetic particles for detection of 8-hydroxy-2´-deoxyguanosine in urine of prostate cancer patients. Electrophoresis, 32, 3207-3220.

Major technology /research activities Dosing machine for screen printed electrodes Real device 3-D Model Controlr unit Electrode reservoir waste Sample imput Cable to potentiostat 3D printing technology

Major technology /research activities Design and manufacturing of Flow injection analysis device for mixing using clever magnetic field coupled with electrochemical detection – technology project Core of electrochemical cell (PDMS) Placement of electromagnetic stirrer Housing of core (ABS). Cell inputs Upper output for preasure regulation in the cell output Cell volume 2000 (ul)

Major technology /research activities Process of manufacturring of prototype… Core of electrochemical cell (PDMS) Positive template (ABS) for negative template from low melting point metal (melting point 75°C) Housing of core (ABS)

Major technology /research activities Process of manufacturring of prototype… Negative template (Wood´s metal - Bi/Pb/Cd/Sn) Final product (PDMS)

Reconstruction of technology part Plan of laboratory of technology after reconstruction EXP. stůl EXP 2 EXP 1 EXP 4 EXP 5 EXP 3 Mechanické úpravy Mechanické obrábění soustruh Mechanické obrábění fréza Návrhy konstrukci a programovaní + pracovní místo Pájecí stanice A testování Osazování pcb optika Pracovní místo u exp stolu + záloha 3D tiskárna Bruska Ovládání 3D tiskárny + pracovní místo (Vstup) (Vstup)

…to all colleagues from the Department of chemistry and biochemistry Acknowledgements …to all colleagues from the Laboratory of biomarkers and flow techniques …to all colleagues from the Department of chemistry and biochemistry