I am sectioning fish (rainbow trout) brains at 16 microns. The brains were kept in PF for 24 hours; washed with Sorenson AB and were kept in Sucrose solution for 24 hours. After this process, they were put on cryomoulds and tissuetek was added and stored at -80 degrees.
The problem i am facing is:
1) my thickness varies from slide to slide. It looks like it varies between 14-18 microns.
2) Question: does temperature plays an imp. role in tissue sections?
3) What should be the ideal temperature for 16 microns slide, considering the fact that the brains were in sucrose solution for 24 hours.