X-Message-Number: 5013 Date: Wed, 18 Oct 1995 14:58:46 -0700 (PDT) From: Doug Skrecky <> Subject: request for help - third answer Suggestion #5: If alcoholic dehydration is used consider adding glycerol to the alcohol based substitution medium. This has had a remarkable effect on preventing fat from leaching from tissue. (J Anat 295-297 Vol.182 1993) Adding polyethylene glycol or acacia gum also seem to help, but these diffuse poorly into tissue. (Am J Clin Pathol 620-623 Vol.88 1987 & J Microscopy 69-84 Vol.92 Pt.2 1970) After 24 years of storage tissue preserved in formaldehyde retained cholesterol, cerebrosides, sulphatides, sphingomyelin and phosphoinositides. Lecithin, phosphatidyethanolamine and phosphatidylserine are lost. (J of Histochemistry & Cytochemistry 704-709 Vol.10 1962) This difference in stability is reflected in the results obtained with formaldehyde treated tissue that is subjected to a methanol/chloroform wash. Even with lipid stabilizers such calcium chloride or uranyl nitrate lecithin and phosphatidylethanolamine show marked losses, while sulphatides and spingomyelin cerebroside are retained. (Histochemical J 323-360 Vol.1 1969) Glutaraldehyde has little effect on retention of lipids, while osmium tetroxide while affording nearly complete retention diffuses poorly. (J Microscopy 1-15 Vol.133 Pt.1 1984) Suggestion #6: Consider using a silica gel bead desiccant packed anhydrous solution of ethylene glycol and glycerol. VS11 (6 M ethylene glycol, 1.8 M glycerol) has a very high glass forming ability. (J Reproduction & Fertility 65-70 Vol.99 1993) By themselves ethylene glycol, glycerol (and triethylene glycol) have been used to dehydrate tissue with great success. (Lab Invest 655-663 Vol.62 No.5 1990 & J Microscopy 195-203 Vol.172 Pt.3 1993) Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=5013