By Norman H. Crowhurst
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This technique is applicable to a wide variety of tissues. A typical PLT dehydration procedure for such tissues, with ethanol, is shown in Table I. The times suggested in the scheme should be modified according to the density and size of the tissue blocks. Other dehydrating liquids require modification of both temperature and concentrations. Freezing points in relation to concentrations for a few of these dehydrating liquids are shown in Fig. 2. TABLE I Dehydration scheme for the PLT technique, with ethanol as dehydrating agent.
L. A. Methacrylates, Pros and Cons The resins are based on methacrylic and acrylic esters due to their low freezing points, low viscosity and their ability to be polymerized by longwave UV irradiation (360 nm) at low temperature. min. temp, Chemical Fixation Rapid Freezing Freeze Substitution 180 K T Low. temp. Vacuum infiltration Low temp. Chemical polymerisation FIG. 1. table) Low temp. v polymerisation ~ 290 K Chemical polymerisation ^ 330 K K11M HM20 K HM23 225 showing the versatility of Lowicryl low temperature resins.
At least three factors have to be considered as essential to minimize loss of structure and biological activity. First, the constituents of the sample should be immobilized by an "inert" procedure, best achieved by rapid freezing, as done in freeze-substitution (Fernandes-Moran, 1959) and freeze-drying, or by chemical fixation with aldehydes. Second, the aqueous environment has to be replaced by a solvent and/or a polymerizable liquid resin, which ideally should have the same properties as water.