An excursion through the ultrastructural world of quick-frozen pancreatic islets

• Published in: American journal of anatomy Vol. 175; no. 2-3; p. 217
• Main Authors: Dudek, R W, Boyne, A F
• Format: Journal Article
• Language: English
• Published: United States 01-02-1986
• Subjects: Animals; Cell Compartmentation; Fixatives; Freeze Drying; Freezing; Glucagon - metabolism; Immunoenzyme Techniques; Insulin - metabolism; Islets of Langerhans - metabolism; Islets of Langerhans - ultrastructure; Male; Microscopy, Electron - methods; Rats
• ISSN: 0002-9106
• DOI: 10.1002/aja.1001750209

In this article we have presented a philosophical and historical perspective on quick freezing, freeze-drying, freeze-substitution, and immunocytochemical localization of pancreatic islet hormones. A compilation of our findings indicates that quick-freezing does not produce any gross distortion of islet tissue; the amount of usable islet tissue for ultrastructural analysis is approximately 13 micron deep from the frozen edge; three different cell types can be identified in quick-frozen tissue based on general morphological characteristics; freeze-substitution with tetrahydrofuran produces a unique ultrastructural appearance in which ribosomes are particularly striking; with the use of protein A-gold, insulin and glucagon can be localized immunocytochemically on silver-gray (50-nm-thick) sections treated with 1% ovalbumin at room temperature overnight; secretory granules of quick-frozen alpha and beta cells may exist in either a swollen or condensed state; swollen beta cell secretory granules contain a filamentous material that demonstrates immunogold labeling for insulin; insulin and glucagon can be localized within the cisternae of endoplasmic reticulum; our methods provide not only discrete immunocytochemical localization of hormone, but also well-preserved cellular compartments; energy electron loss spectroscopy (EELS) has shown that quantifiable nitrogen maps can be used as an index of hormone packaging in secretory granules; and the sectioning properties of secretory granules at the ultramicrotome change when islet tissue is unosmicated and sectioned on glycerol.