Freeze-substitution and the preservation of diffusible ions.
Freeze-substitution of biological material in pure acetone followed by low-temperature embedding in the Lowicryls K11M and HM23 yields stable preparations well suited for sectioning and subsequent morphological and microanalytical studies. Transmission electron microscopy of dry-cut sections shows that diffusible cellular thallium ions (T1+) of T1(+)-loaded muscle are localized at similar protein sites in freeze-substituted as in frozen-hydrated preparations. A comparison of X-ray microanalytical data obtained from freeze-dried cryosections and sections of freeze-substituted normal (potassium-containing) muscle shows that K+ ion retention in the freeze-substituted sample is highly dependent on the freeze-substitution procedure used so far, in the best case, about 67% of the cellular K+ is retained after freeze-substitution in pure acetone and low-temperature embedding. It is concluded that the macromolecules during the preparative steps and that ion retention may be increased by further optimizing freeze-substitution and low-temperature embedding.
Edelmann
L
L
Universität des Saarlandes, Homburg/Saar, Germany.
D000096:Acetone; D000818:Animals; D015925:Cryopreservation; D004058:Diffusion; D004577:Electron Probe Microanalysis / Q000379:methods*; D005612:Freeze Drying; D005629:Frozen Sections; D008854:Microscopy, Electron / Q000379:methods*; D009132:Muscles / Q000737:chemistry / Q000648:ultrastructure*; D011188:Potassium / Q000378:metabolism; D011894:Rana pipiens; D013793:Thallium / Q000378:metabolism
D000096:Acetone; D013793:Thallium; D011188:Potassium