This thesis aims to assess the possibility of the usage of various biological fixatives for fish cell and tissues samples in order to extend its storage for later flow cytometric measurement of DNA content. The model species chosen were sterlet and tench, from which three types of samples were obtained: blood and fin tissue of subadult / adult individuals and tail tissue of hatched larvae. Altogether 13 fixation methods were tested for each type of sample of both model species. Methods were chosen based upon their easy feasibility and low time-consumption. The samples were measured on flow cytometer in native state immediately after sampling and placing in physiological saline and after 1, 5 and 10 days of fixation during which they were stored in a fridge or in a freezer at -80 ?C. Their analysis was carried out simultaneously with standards native cells from tench fin tissue when investigating sterlet samples, and commercially available fixed trout erythrocytes for tench samples. A fluorochrome used was 4',6-diamidine-2'-phenylindole dihydrochloride (DAPI; with excitation/emission maxima 358 / 461 nm). Based on the evaluation of coefficients of variation (CV) of fixed samples and the changes in their fluorescence levels in comparison with native state, optimal procedures for extended storage of all types of samples from both model species are suggested.
| Identifer | oai:union.ndltd.org:nusl.cz/oai:invenio.nusl.cz:375884 |
| Date | January 2018 |
| Creators | HUBÁLEK, Martin |
| Source Sets | Czech ETDs |
| Language | Czech |
| Detected Language | English |
| Type | info:eu-repo/semantics/masterThesis |
| Rights | info:eu-repo/semantics/restrictedAccess |
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