3D Volume Serial Block Face Imaging Facility. (360G-Wellcome-108461_Z_15_Z)

Compartmentalization of mutually exclusive reactions in different regions of cells by membrane-enclosed organelles or by self-assembling macromolecular complexes is a critical mechanism of life. The fluorescent tags of optogenetics can display proteins confined to compartments in living cells, but gives no glimpse of the underlying ultrastructure. The electron microscope has the resolution to display cellular ultrastructure, but within very thin slices (100nm depth) from which it is difficult t o understand, measure and compare the complex cellular ultrastructure, particularly with elaborately interdigitating cells in vivo such as stem cells or neurons. Serial Block Face Scanning Electron Microscopy (SBFSEM) provides ultrastructural resolution thoughout a much longer ( m to mm) sample depth, enabling quantitative analysis of ultrastructural features throughout the length of most complex cells. Kings seeks support from The Wellcome Trust to establish a SBFSEM with high resolution ( < 3nm), optimized for biological samples, to enable quantitative analysis of cellular ultrastructure in three-dimensions. The information from this third-dimension will transform many basic and clinical science applications at Kings and with our collaborators. It will also be pivotal in the project at Kings, involving Nikon and JEOL, in identifying optogenetic fluorochromes within their 3D-ultrastructural context in the SBFSEM.