Gregory Hirsch — Department of Energy SBIR Phase II: C53-27a

Description

C53-27a-271143Transformative advances in cryogenic electron microscopy (cryo-EM) have expanded our ability to visualize cellular and biomolecular structure, often at atomic resolution. However, a significant hurdle for achieving the full potential of cryo-EM, including work performed at facilities associated with DOE, involves difficulties with sample preparation. Reliably producing vitrified ultrathin samples on grids, especially those associated with single particle analysis (SPA), is very challenging. Too often, grids delivered to these facilities are found inadequate for collecting good data, and the investigator’s precious user time on microscopes is wasted. This project is addressing these difficulties by developing a novel tool to rapidly screen cryo-EM grids at point- of-origin by providing critical information on the thickness of the vitreous ice/sample layer. In addition to identifying the best sample grids for further cryo-EM analysis, this information can enable near real-time optimization of vitrification procedures to increase sample preparation workflow. The technology being developed could be used in either standalone instruments or incorporated into full robotic vitrification systems. A prototype instrument was designed and constructed to validate our approach for rapid and precise screening of cryo-EM sample grids. Its initial performance was demonstrated by imaging test grids at cryogenic temperature. Lateral spatial resolution was observed to be in the target range below 1?m. Several different embodiments of the tool were evaluated for incorporation with the more sophisticated Phase II prototype instrument. IP protection for this technology was started, and strategic relationships for Phase II were set up. Significant improvements to the instrument and its performance will be developed and incorporated into the prototype. This instrument will be used in conjunction with work at the Stanford-SLAC Cryo-EM Center to validate its utility in a real research environment. Efforts to commercialize this technology will be pursued. Cryo-EM has experienced explosive growth in recent years, with projected worldwide annual revenue for just microscopes to exceed $1B before 2030. Improved sample preparation equipment represents a substantial percentage of this market sector. Cryo-EM data has become central for many areas of basic and applied science, including drug discovery in the huge biotech and pharma industries. This project seeks to develop a crucial tool for greatly improving the workflow for cryo-EM analysis in many disciplines of life sciences, medicine, environmental research, and other fields requiring structure determination at the highest possible resolution.