Oolong tea extract as a substitute for uranyl acetate in staining of ultrathin sections based on examples of animal tissues for transmission electron microscopy

Abstract

Oolong tea extract (OTE) was assessed for its potential as an electron stain to substitute for uranyl acetate (UA) in transmission electron microscopy (TEM). A comparative analysis of the ultrastructures of biological specimens (i.e. compound eye and sciatic nerve tissue) stained by different methods (UA and 0.1% OTE) has been performed. Results revealed that there was no significant difference between the OTE double staining method and the traditional double staining method, except the contrast in the OTE method was slightly lower than the UA method. Nevertheless, the OTE method yielded better or equal details in collagen fibres, neurofilaments and basal lamina in the mammalian sciatic nerve samples, as well as in the microvilli between the cornea and crystalline cone of the insect compound eye. On the whole, it was suggested that OTE could potentially be used as a nonradioactive and hazard-free alternative to UA in TEM staining.

Oolong tea and LR‐White resin: a new method of plant sample preparation for transmission electron microscopy

“…water or PBS) did not affect the quality of TEM results (data not shown), which is consistent with the observations by Carpentier et al . () This may be because of the different types of samples, since the samples in Carpentier's () and our studies were derived from plant tissues, whereas in Sato's () and our previous works (He & Liu, , ), the samples were of animal or bacterial origins. But it is noteworthy that a freshly prepared OTE staining solution is essential to achieve successful staining, as is confirmed in all of our studies.…”

“…This reduced the contrast between cytoplasm and organelles. However, 0.1% OTE previously demonstrated better performance than 0.05% OTE in bacterial cells and animal tissues ultrastructure observations (He & Liu, , ). This difference could be due to different electron adsorption capacities in different samples.…”

“…OsO 4 acted not only as a fixative, but also as an electron blocking staining reagent. Although post fixation with OsO 4 was essential for high‐contrast staining, staining only with OsO 4 did not produce a sufficiently high contrast to satisfactory TEM observations (He & Liu, ). The use of single staining solution alone (UA or OTE) also did not produce sufficient increase in contrast of the given samples (He & Liu, ) (Fig.…”

“…Previous studies demonstrated that the nature of the staining solution could affect the staining effect in ultrathin sections. In animal tissues (Sato et al ., ; He & Liu, ) and bacterial cells (He & Liu, ), OTE was dissolved in a specific buffer. In this research, however, the nature of the OTE solvents (i.e.…”

“…Aiming to solve these three problems (optimizing sample preparation protocols, shortening the time needed for specimen preparation, searching safer staining solution for replacing UA as a contrasting agent), we have previously performed assays that combined LR‐White resin with the use of polyphenolic compounds (Oolong tea extract, OTE) for bacteria and animal tissues (He & Liu, , ). In this study, this approach was extended to plant specimens.…”