Imaging of subcellular structures by scanning ion microscopy and mass spectrometry. Advantage of cryofixation and freeze substitution procedure over chemical preparation

摘要: With the IMS 4F, a scanning ion microscope and mass spectrometer (SIMS), it is possible to map chemical elements with lateral resolution of about 250 nm over field view 50 × μm2. Such conditions should enable imaging subcellular structures constitutive ionic species such as CN−, P−, S−. The study was performed on heart renal tissues prepared either by procedure or cryofixation-freeze substitution (CF-FS) prior embedding. Heart tissue chosen because cardiocytes display simple structural organization whereas kidney tubular cells more complex. Whatever preparation procedure, nuclei were easily identified due their high P− content. S− images obtained showed weak contrasts inside cytoplasm so that difficult recognize organelles. After CF-FS, enhanced contrasted allow organelle (mitochondria, myofibrils, lysosomes, vacuoles, basal lamina, etc) characterization. This work demonstrated CF-FS suitable than method reveal composition. improvements in these an essential step establish correlation between localization trace element (or molecule tagged isotopes particular atoms) its targets.