Comparative study of hydrophilic and hydrophobic ionic liquids for observing cultured human cells by scanning electron microscopy

Abstract

An ionic liquid (IL) is a salt that remains in the liquid state at room temperature. It does not vaporize under vacuum and imparts electrical conductivity to samples for observation by scanning electron microscopy (SEM). Recently, the usefulness of ILs has been widely recognized. In our previous study, one of the ILs 1-ethyl-3-methylimidazolium tetrafluoroborate (EtMelm(+) BF(4)(-)) was used for SEM analysis of biological samples. In comparison with the conventional method, samples prepared using EtMelm(+) BF(4)(-) provided more detailed SEM images of the cell ultrastructure, enabling the observation of protrusions. In addition, the IL treatment is a less time consuming and simple method that does not include dehydration, drying, and conductivity treatments, which are an essential parts of the conventional method. In this study, we compared the usefulness of four hydrophobic and three hydrophilic ILs for SEM to observe fixed cultured human A549 cells. All ILs worked well to prevent "charge-up" effect for SEM observation. However, the hydrophilic ILs tended to provide clearer images than the hydrophobic ILs. We concluded that various ILs can be used for SEM sample preparation and their application to a wide range of fields is anticipated in future.

Electron microscopy of Staphylococcus epidermidis fibril and biofilm formation using image-enhancing ionic liquid

“…These FE-SEM results indicate that fine morphologies can be clearly observed using hydrophilic IL without any conductive coating. Ishizaki et al have reported that a simple technique using hydrophilic IL enables the observation of the fine thread-like morphologies of cultured human cells [28]. In good agreement with this report, we observed fine morphologies of filamentous fibrils using an IL.…”

“…On the other hand, the observation of biological materials by electron microscopy without structural deformation is a real challenge. In this regard, combining ILs with biological materials can be the most simple and attractive approach to reveal the morphological structure by electron microscopy [27,28]. We found that the osmotic pressure of biological materials needs to be maintained to observe the fine morphology [29].…”

Immuno‐detection of mRNA‐binding protein complex in human cells under transmission electron microscopy

“…HeLa and A549 are most popularly used and well documented cell lines, from our previous studies, we have clearly understood the biogenesis of RBM8A and Upf2 in HeLa and A549 (Ishigaki et al, ; Ishigaki et al, ; Ishigaki, Nakamura, Tatsuno, Ma, & Tomosugi, ; Tatsuno & Ishigaki, ; Tatsuno, Nakamura, Ma, Tomosugi & Ishigaki, ). We performed immunostaining for Y14 and Upf2 in an ultrathin section for TEM as shown in Figure .…”

A Facile Method for Simultaneous Visualization of Wet Cells and Nanostructured Biomaterials in SEM using Ionic Liquids

“…[17] Moreover, RTIL treatment can mitigate artifacts in biological samples such as wrinkling, shrinkage, and cracking, and it may provide a closer morphological representation of the natural hydrated microbe state. [17] Researchers have utilized RTILs purely as an alternative to metal sputter coating, [17][18][19][20] while others have used RTIL to image living bacterial cells and red blood cells with SEM. [21,22] While these studies have shown the capacity to image cells, they have often been limited to single endpoint studies on non-complex substrates with simple organisms such as bacteria.…”