Assembly for Cryo-SEM/Raman Microspectroscopy sample analysis

0568569 - ÚPT 2023 RIV CZ eng A - Abstract
Láznička, Tomáš - Hrubanová, Kamila - Ježek, Jan - Kizovský, Martin - Bernatová, Silvie - Mrázová, Kateřina - Obruča, S. - Zemánek, Pavel - Krzyžánek, Vladislav
Assembly for Cryo-SEM/Raman Microspectroscopy sample analysis.
16th Multinational Congress on Microscopy, 16MCM, 04-09 September 2022, Brno, Czech Republic. Book of abstracts. Brno: Czechoslovak Microscopy Society, 2022 - (Krzyžánek, V.; Hrubanová, K.; Hozák, P.; Müllerová, I.; Šlouf, M.). s. 63-64. ISBN 978-80-11-02253-2.
[Multinational Congress on Microscopy /16./. 04.09.2022-09.09.2022, Brno]
R&D Projects: GA TA ČR(CZ) TN01000008
Grant - others:AV ČR(CZ) MSM100652102
Program: Program na podporu mezinárodní spolupráce začínajících výzkumných pracovníků
Institutional support: RVO:68081731
Keywords : Cryo-SEM * Raman spectroscopy * electron microscopy
OECD category: Electrical and electronic engineering
https://www.16mcm.cz/wp-content/uploads/2022/09/16MCM-abstract-book.pdf

Correlative imaging using different methods of observation is now becoming more and more popular. It allows the interconnection of various information of the sample accessible only by
individual techniques. It is crucial to find and observe the same place on the sample. The same objects and their locations (cell shape and arrangement) can be correlated with each other,
especially with software when 2 different imaging devices were used. Combination of the Cryo-Scanning Electron Microscopy (Cryo-SEM) and Cryo-Raman Micro-spectroscopy (Cryo-Raman) gives many benefits. Surface, structure or morphology of the sample can by studied using Cryo-SEM, and detection and identification of elements or compounds in the examined sample using Cryo-Raman. For observation of various, especially biological samples, their freezing to the temperature of liquid nitrogen is used. The motivation was to design an assembly for motion and observation using Cryo-Raman and its connection with commercial holders used for the observation of vitrified samples by Cryo-SEM. Assembly allows movement of the sample to a known position and measuring the cryogen level using LabVIEW software. Our assembly consists of a Dewar vessel, which contains liquid nitrogen. The stage is inside the Dewar vessel below the liquid level and is made by 3D printing and enables 3 positions for individual types of sample holders (Leica microsystems, Gatan and Quorum technologies). The Dewar vessel lies on piezoelectric stage allowing movement in both X and Y direction. The stage is connected to the Raman spectrometer table providing displacement in the Z direction. The Dewar vessel is covered with Plexiglas to minimize contamination. Assembly was tested on dried samples and polystyrene in cryo temperatures with known spectrum to verify functionality. Then biological samples including C. necator H16 were tested. System allows more efficient correlation between individual analyses of microorganisms containing biopolymer particles. For example, several representatives of soil microorganisms, such as bacteria C. necator H16 which are able to produce polyhydroxyalkanoates (PHA). PHAs are polyesters of hydroxyalkanoic acids, which serve as energy storage. PHAs attract attention as an alternative to petrochemical plastics, which can be biotechnologically produced from waste streams of some industrial processes. It was proved that the combination of Raman spectroscopy with the cryo-SEM technique can provide a deeper insight into the chemical and mechanical properties of polymeric granules inside the bacterial cells and it will serve for future research.
Permanent Link: https://hdl.handle.net/11104/0339859