Engineering Arts (EA) proposes the development of a detachable nozzle piezoelectric dispense system for use in Cryo-TEM (Cold - Transmission Electron Microscopy) sample preparation instrumentation. Cryo-TEM is an important tool for studying the structure of biological macromolecules in their native aqueous state. The challenging and laborious process of preparing 'thin-ice'frozen aqueous samples for Cryo-TEM has remained essentially unchanged for decades. Typically a single aqueous biological sample (~3?l) is manually applied onto a thin porous Electron Microscopy (EM) grid which is then blotted to remove excess sample and then plunged into liquid-ethane for flash freezing. This process hopefully leaves enough regions on the grid with the proper vitrified 'thin-ice', typically 50 to 200nm thickness, required for EM imaging. The detachable piezoelectric nozzle dispense system proposed here is a key innovation that will lead to productivity, quality and capability breakthroughs for Cryo-TEM workflow. The following key benifits for Cryo-TEM process will be realized: 1. Apply many different samples (or sample conditions) to each Cryo-TEM grid. 2. Increase process control and yields of acceptable thin-ice over current standard blotting methods. 3. Lower starting sample volume requirements and consumption per grid. 4. Allow time-resolved study of mixing / binding events of multiple samples. EA will design, fabricate and optimize a multichannel detachable nozzle piezoelectric dispense system that is capable of dispensing droplets (less than 50 picoliters) of several different biological samples (or sample conditions) onto each TEM grid. A three channel detachable nozzle piezoelectric dispense system retrofitted onto a custom Cryo-TEM sample preparation instrument (Spotiton instrument platform) at NRAMM (National Resource for Automated Molecular Microscopy, Scripps, La Jolla CA.) for testing, optimization and validation.
In this grant, Engineering Arts proposes development of a tool that will allow scientists to more easily construct high resolution molecular models of proteins, viruses and other biological structures. Understanding life processes at the molecular level forms scientific foundations for understanding disease leading to potential diagnostics and therapies including new drugs.