Schellens J P, Vreeling-Sindelárová H, Van den Munckhof R J, Frederiks W M
Department of Cell Biology and Histology, Academic Medical Centre, University of Amsterdam, The Netherlands.
Histochem J. 1995 Aug;27(8):609-14.
Glycogen phosphorylase activity has been demonstrated at the ultrastructural level in liver and heart tissue of fasted rats. Unfixed cryostat sections were incubated by mounting them on a semipermeable membrane stretched over a gelled incubation medium. The medium contained a high concentration of glucose 1-phosphate which enables indirect detection of glycogen phosphorylase activity on the basis of the synthesis of glycogen. Tissue fixation, dehydration and embedding for electron microscopical study were performed after the incubation had been completed. The ultrastructure of both liver and heart tissue was rather well preserved. Glycogen granules resulting from glycogen phosphorylase activity were found in the cytoplasmic matrix of both hepatocytes and cardiomyocytes; no relationship with membranous structures could be detected. It is concluded that the semipermeable membrane method is well suited for localizing cytosolic enzyme activities at the ultrastructural level without prior tissue fixation; this opens further perspectives for correlations between histochemical and biochemical data.
在禁食大鼠的肝脏和心脏组织中，已在超微结构水平上证实了糖原磷酸化酶的活性。将未固定的低温恒温器切片安装在覆盖于凝胶化孵育介质上的半透膜上进行孵育。该介质含有高浓度的葡萄糖-1-磷酸，这使得能够基于糖原的合成间接检测糖原磷酸化酶的活性。孵育完成后，进行用于电子显微镜研究的组织固定、脱水和包埋。肝脏和心脏组织的超微结构保存得相当完好。在肝细胞和心肌细胞的细胞质基质中均发现了由糖原磷酸化酶活性产生的糖原颗粒；未检测到与膜结构的关系。得出的结论是，半透膜方法非常适合在不预先进行组织固定的情况下在超微结构水平上定位胞质酶活性；这为组织化学和生化数据之间的相关性开辟了进一步的前景。
