Design, construction and testing of a new shock freezing system during parabolic flight

Background: Fixation of tissue is a common method to stop biological processes and preserve a status of interest for later examination. Popular done with liquid nitrogen, this is not possible during parabolic flight investigations, due to safety issues. However, for the investigation of the direct effects of gravity, respectively the absence of it, it is necessary to fixate the tissue during or right after the experience of weightlessness. Hence the aim of this thesis was to invent and establish a method for shock freezing biological samples during parabolic flight. A carbon dioxide based freezing system was designed and constructed. Tissue was frozen in a steel tube where dry ice was created from carbon dioxide out of two gas cylinders conducted over a pressure system. The method was tested on Arabidopsis thaliana seedlings and compared to known freezing techniques, like chemical fixation and liquid nitrogen. In a comparison of quality and quantity the RNA of deep frozen material was analyzed. Results: A system was constructed where samples are deep frozen by dry ice in a steel cylinder. Gaseous carbon dioxide from a gas cylinder is conducted over a pressure line into a steel cylinder, where dry ice is formed due to a pressure gradient. Arising gas is conducted out of the chamber by a pressure system. The whole system is fixed on a rack construction, which can be mounted on the air craft. The quality of the new system was found to be comparable to other methods. Analyzed RNA was found to be of good quality. Time until shock freezing could be reduced to under a minute and duration of freezing could be guaranteed long enough for a whole flight day during a parabolic flight campaign. Conclusion: The Pre-testing on ground gives evidence that the system can be used during in flight configuration. The system is now to be used in the 10th parbolic flight campaign of the DLR for a first operational test. In future campaigns it will be available to different research groups as fixation method.Validerat; 20101217 (root