Labeling of Ultrathin Resin Sections for Correlative Light and Electron Microscopy

摘要: Correlative microscopy combines the versatility of light microscope with excellent spatial resolution electron microscope. Here, we describe fast and simple methods for correlative immunofluorescence immunogold labeling on very same ultrathin section. The protocols are demonstrated sections tissue samples embedded in methacrylate Lowicryl K4M. Ultrathin mounted (EM) grids stained simultaneously fluorescent gold markers. For detection primary antibodies, applied either protein A or immunoglobulin G (IgG) combination secondary antibodies coupled to Alexa488 Alexa555. Alternatively, marker FluoroNanogold was used, followed by silver enhancement. have be analyzed first at then transmission (TEM), because fluorescence is bleached beam. Labeled structures selected can identified TEM high resolution. This way, signals directly correlated corresponding subcellular area interest.