Cryofracture of paraffin‐embedded heart muscle cells

Abstract

SUMMARY A comparative study of internal cellular structures of the sheep ventricular myocardium has been conducted by scanning electron microscopy (SEM) and by transmission electron microscopy (TEM). Access to the cell interior for three‐dimensional studies was obtained by cryo‐fracturing paraffin‐embedded tissue frozen in liquid nitrogen. For accurate localization of structures of special interest thick paraffin sections were examined in the light microscope (LM). Based on the information gained, it was possible to fracture the block in a desired plane. The fracturing was carried out by a light blow to a precooled scalpel held against the surface of the block, which was immersed in liquid nitrogen. After thawing and deparaffinizing at room temperature in several baths of xylene, the tissue pieces were critical point dried using CO2. As xylene was found to be miscible with CO2, it also served as an intermediate fluid. This method resulted in good preservation of the myofibrils, mitochondria, sarcoplasmic reticulum and transverse tubules (T‐tubules), which was confirmed by TEM studies of conventionally prepared tissue and of tissue originally prepared for SEM.

An ultrastructural study of cryofractured myocardial cells with special attention to the relationship between mitochondria and sarcoplasmic reticulum

“…Similar structures were also exposed in thc quenchfrozen hydrated tissue, where the ice formation had resulted in an extensive separation of the cell organelles. In the past such structures have been observed in scanning electron micrographs from the myocardium of dog, rat, lamb, sheep and pigeon (Sybers & Ashraf, 1974, 197%;Sheldon et al, 1976;Dalen et al, 1978;Myklebust er al., 1980). The present transmission electron micrographs of critical point dried material confirmed that these structures are part of the SR.…”

“…The various cryofracturing procedures have been reviewed by Dalen (1983). Several authors have applied such techniques for exposing the myocardial cell interior for morphological studies in the scanning electron microscope (SEM) (Haggis, 1970;Sybers & Ashraf, 1973, 1975a, 1975McCallister et al, 1974;Dalen et al, 1978;Myklebust et al, 1980).…”

Effect of cooking temperatures on characteristics and microstructure of camel meat emulsion sausages

“…Scanning electron microscopy (SEM) examination was conducted for prepared raw batter and cooked emulsion sausage . One core 1 × 1 × 1 cm from each sample was fixed in 2.5% glutaraldehyde for 2 h and trimmed to less than 2 mm thickness, then washed three times with 0.1 mol L −1 phosphate buffered saline each for 10 min.…”

A new technique for visualization of internal structure by SEM

“…(1974) and Dalen et a/. (1978) have described a procedure in which paraffin embedded tissue was frozen in liquid nitrogen, deparaffinized and dried.…”