CRYOFIXATION, CRYOSUBSTITUTION, AND IMMUNOELECTRON MICROSCOPY - POTENTIAL ROLE IN DIAGNOSTIC PATHOLOGY

In immunoelectron microscopic investigations, retention of antigenic s ites is crucial. Methods for preparing samples for conventional electr on microscopy involve chemical fixatives followed by dehydration in or ganic solvents and embedding in plastic resins, all procedures potenti ally detrimental to antigenicity. Cryomethods provide a physical fixat ion alternative for the preparation of biological samples for ultrastr uctural, immunocytochemical, and microanalysis studies without the use of any chemicals. This can be particularly useful in diagnostic patho logy, providing an alternative to conventional fixation methods which sometimes destroy the antigen in question. The recent development of a portable cryofixation device, the PS1000 Portable Metal Mirror Ultra- Rapid Cryofixation Unit (Delaware Diamond Knives, Inc., Wilmington, DE , USA), provides an opportunity to freeze tissue immediately after pro curement for use in diagnostic immunocytochemistry studies. This feasi bility study examined the quality of tissue preservation with this dev ice, in terms of both preservation of cellular ultrastructure and immu nolabeling. Human tonsil and thymus tissue was slam frozen and, after cryosubstitution in Lowicryl K11M, was examined by immunoelectron micr oscopy. Good ultrastructural preservation was obtained and reasonable immunolabeling with antibodies to AE1/AE3 keratin filaments was also o bserved.