As part of an established core facility, we request an OMX Blaze structured-illumination microscope (SIM) from Applied Precision. Key cellular processes occur in structures and assemblies smaller than resolvable by confocal microscopy, but still requiring the ease and specificity of fluorescence labeling. Addressing multiple experimental systems, our super-resolution needs have five constraints. (1) The volumes we must explore are often deep within cells and tissues, significantly distant from the coverslip. (2) All studies require axial super-resolution, as well as lateral super-resolution. (3) The relevant structures, spanning large protein assemblies to organelles to the nucleus, are typically macromolecular assemblies 100 nm or larger. (4) The assemblies often interact with many other molecules, requiring a robust palette of colors for simultaneous visualization. (5) For some live-cell dynamics, speed of imaging is important. After extensive testing of multiple technologies, only SIM is appropriate for our needs. Furthermore, we have preliminary SIM data for all Users, thoroughly exploring each vendor's implementation of SIM. Compared to existing equipment anywhere at Johns Hopkins, the proposed instrument will provide an order-of-magnitude superior performance that will enable key NIH-funded projects.
| Yang, J-Ming; Bhattacharya, Sayak; West-Foyle, Hoku et al. (2018) Integrating chemical and mechanical signals through dynamic coupling between cellular protrusions and pulsed ERK activation. Nat Commun 9:4673 |
