X-Message-Number: 5193 Date: Thu, 16 Nov 1995 16:17:56 MST From: "Richard Schroeppel" <> Subject: damage assessment & ice microscopy One of our constant arguments is "How much freezing damage ...?" We should be investigating ways to examine frozen tissue, without thawing it first. The only technique I know of is freeze fracture; I've no clue as to its applicability. There are various heroic techniques for examining a few atoms of a surface in a vacuum; these are probably unsuitable for our needs, since they don't look at enough material, and we're not ready to use atomic level data anyway. One possibility would be to develop a good method of slicing a block of frozen tissue without melting the surface we are trying to examine. The top millimeter or so could be examined with an ordinary microscope. Obvious candidates are lasers, sharp knives, and corrosives; whether they could produce an interpretable surface is questionable, but worth looking into. We might also see what techniques have been used by other scientists who were reluctant to thaw their subjects - ice-man, mammoths, ice-cores. Another possibility would be to prestain the test animal (before freezing) - as an example, if we knew that fluorescein didn't enter cells, we could give the animal fluorescein, and see what the distribution looked like after the freezing protocol. Conceivably this could be combined with NMR to develop high resolution data. An outside shot would be to look for a sequence of fluid substitutions that preserved tissue & cell structure. It's already been mentioned that ice dissolves readily in methanol. The problem to be overcome is to find a sequence of substitutions that leaves some tissue to examine. As always, a bad preservation result doesn't mean a lot, while a good preservation result would validate the freezing protocol. Development of methods for examining frozen tissue would probably have other scientific uses besides cryonics. Rich Schroeppel Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=5193