Study on the Methods of In Situ Embedding Monolayer Cultured Neurons for Electron Microscopy

Abstract: To investigate an optimized in situ embedding method of monolayer cultured neurons， the rat cortical neuronal cells were used as the experimental samples to be embedded in resin by three methods，respectively. The results showed that the conventional cell suspension-centrifugation sample preparation was easy to cause mechanical damage and hard to remain cells in situ. The cells cultured on the glass coverslips kept their original position unchanged， however their ultrathin sections were easily broken and readily dissolved in the water.Also， these cells always exhibited overall weak contrast and incomplete structure under the transmission electron microscopy (TEM). When neuronal cells were cultured on Thermanox coverslips， the connections between adjacent cells could be easily identified. The ultrastructures of these cells were much clearer and their overall contrast was better than that of the cells cultured on the glass coverslip. The process of in situ embedding monolayer cell on the plastic film using in the third method was more simple and easily carried out， compared with that using in the former two methods. Taken together， these results provides new insights into and useful methods for researching neuronal cell ultrastructure， that will greatly help to develop novel techniques combing three-dimensional reconstruction and correlative light and electron microscopy (CLEM) simultaneously.

CSTR: 32200.14.cjcb.2016.12.0010