Preparation of cellular samples using graphene cover and air-plasma treatment for time-of-flight secondary ion mass spectrometry imaging

We report on sample preparation methods based on plasma treatment for an improvement of multiple molecular ion images of cellular membranes in the ToF-SIMS method. The air-plasma treatment of fixed cellular samples efficiently removed the organic residues of any solutions used during sample preparation and improved the quality of ToF-SIMS images due to the resulting clean surface. We also studied cell preparation methods that combine single-layer graphene covering with air-plasma treatment to achieve a synergistic effect that eliminates background spectra by organic impurities while minimizing morphological cell deformation in a vacuum environmental analysis. When the cellular sample on the glass substrate is completely covered with the single-layer graphene, the cells trapped between the graphene and the substrate can effectively reduce morphological deformation by slow-dehydration. After slow-dehydration of cells is completed inside the graphene-cover, the covered graphene layer can be peeled off by air-plasma treatment, and the unwanted organic residues on the surface of cells and substrate can also be removed by plasma cleaning, thereby much improving ion imaging of cells with the ToF-SIMS method. It is confirmed that the cell samples in which the graphene-cover was removed by air-plasma treatment maintained their morphology well in comparison with the rapid air-dried cells in atomic force microscopy (AFM) and ToF-SIMS images.