My lab is having a problem preparing thin plastic sections (70 nm thick) for standard Transmission Electron Microscopy. We are using a Leica UC6 ultramicrotome. The samples are of brain and retina that are embedded in epoxy resin, either Durcupan or LX112-Araldite (similar to Epon 812-Araldite).
Here is the problem:
We can cut good semithin sections at 0.5- 1 um thickness, but when we switch over to cut thin sections at 70 nm for TEM, the advance control seems to malfunction. When making the switch the first thin section that should be 70 nm, tends to be thicker than that, and subsequent passes of the specimen past the knife edge either do not contact the knife resulting in no section, or will produce shavings from the upper part of the specimen block face, but not the entire block face. After several more passes of the specimen, we may get a full section of the block face but again it is often thicker than 70 nm, and then this sequence of taking one section that is too thick followed by no sections repeats.
The hardware (Knife, specimen holder, etc.) is all tightened well, so this shouldn't be a problem with things being loose.
Has anyone encountered this particular problem before? Any insight or suggestions would be greatly appreciated. Thanks.
Dave Sherry
I get this same problem when the blocks I'm sectioning are too soft, under-polymerised. When this happens, I place the blocks into the 60 degree C oven overnight. This usually corrects the problem.
B
Have you tried adjusting the angle of the blade to the specimen? I have not cut your specific specimens but have had this issue with a microtome after I had it serviced. The service technician had hit a knob that loosened the blade angle. It caused the exact same problem you are seeing. The first section was too thick. Subsequent attempts at sectioning get no slice at all, then followed by a good slice of normal thickness. Once I re-adjusted the blade angle back to suggested specs, problem solved! Good luck!
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