Universität Wien

300104 UE *Hands on" Cryopreparation of Biological Samples for Ultrastructural Studies (2018W)

5.00 ECTS (4.00 SWS), SPL 30 - Biologie
Continuous assessment of course work

Registration/Deregistration

Note: The time of your registration within the registration period has no effect on the allocation of places (no first come, first served).

Details

max. 6 participants
Language: German, English

Lecturers

Classes

Obligatory safety instructions will be given by Daniela Gruber on 5th October 2018, 9 a.m. to 10 a.m. in the Conference Room Zoology. UZA 1, Biozentrum Althanstraße 14, 1090 Vienna, room no. 1.015.

Thursday. 15.11.2018 09:00 – 12:00 lecture SR1, 13:00 – 17:00 Practical work, EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Friday. 16.11.2018 09:00 – 17:00 Practical work, EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Monday, 19.11.2018 09:00 – 17:00 Practical work, EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Tuesday. 20.11.2018 09:00 – 17:00 Practical work, EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Wednesday. 21.11.2018 09:00 – 17:00 Practical work, EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
22nd Nov. - 7th Dec 2018 Personal appointments for cutting TEM sections with an ultramicrotome. EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Monday, 10.12. 2018 Group A: Work with the TEM. EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Tuesday, 11.12. 2018 Group B: Work with the TEM. EM-LAB Core Facility Cell Imaging und Ultrastrukturforschung.
Friday, 14.12.2018, 9:00 – 13:00 Final discussion, presentation of the result (talks). SR 1


Information

Aims, contents and method of the course

The life-science community increasingly includes cryopreparation techniques for microscopic and analytical methods. Ultrarapid cryoimmobilization of living samples in combination with low-temperature dehydration-and fixation (freeze substitution) became a quality standard for the preservation of animal- and plant tissues for transmission electron microscopy that enabled new approaches to tackle biological questions.
The course educates students in the principles of two substantially different freezing techniques, namely, high-pressure freezing and plunge freezing. Both techniques will be used during the course to freeze selected biological samples that are either alive or chemically fixed (as they result frequently from field experiments). The samples will be freeze substituted by using agitation modules inserted in the cryochambers of automated freeze substitution systems (AFS). The application of those modules, developed in our EM lab, allows drastic reduction of the processing time (see literature). Subsequently, the samples will be embedded either in epoxy resin or Lowicryl. The latter is in particular of interest for applications that aim on immunolabeling.
The sectioning of the resin embedded samples will be arranged according to individual requirements for practicing within a separate stage of the course. Practical work will be complemented by lectures and discussions, which will highlight the role of cryopreparation in the concept of biological sciences and its combination with other methods. Finally, the sample quality will be evaluated in the light- and electron microscope. The results will be presented in a short talk.
In consequence, the course provides a preparative base for more complex experimental approaches, which may include phenotyping at ultrastructural level, labeling methods (immunolabeling, in situ hybridization), element analysis (EDX and EELS) and, increasingly, also NanoSIMS.

Assessment and permitted materials

Participation: 40%
Oral presentation of the results: 40% (23th March 2018)
Protocol: 20%
Every item has to be positive.

Minimum requirements and assessment criteria

Theoretical knowledge (lecture). Application of equipment for sample preparation and TEMs (practical work). Evaluation, discussion and interpretation of the results.

Examination topics

Ability to select suitable methods for cryopreparation for TEM samples, practical work in our chemistry lab, save handling of cryogens, liquid nitrogen and propane, handling of equipment for sample preparation incl. plunge freezer, high-pressure freezer, ultrathin sectioning with ultramicrotomes, work with TEMs, interpretation and evaluation of the results.

Reading list

Mielańczyk L, Matysiak N, Klymenko O and Wojnicz R (2015). Transmission Electron Microscopy of Biological Samples. In: The-transmission-electron-microscope-theory-and-applications. IntechOpen. See: https://www.intechopen.com/books/the-transmission-electron-microscope-theory-and-applications/transmission-electron-microscopy-of-biological-samples
Reipert S, Goldammer H, Richardson C, Goldberg MW, Hawkins TJ, Hollergschwandtner E, Kaufmann WA, Antreich S, Stierhof YD. Agitation Modules: Flexible Means to Accelerate Automated Freeze Substitution. J Histochem Cytochem. 2018 Jul 1:22155418786698.
https://www.growkudos.com/publications/10.1369%25252F0022155418786698/reader

Association in the course directory

MBO 7, MGE III-2, MMB W-2, MMEI III, MZO 2, MZO W-1, WZB, M-WZB

Last modified: Fr 31.08.2018 08:43