Ultrafast Biophysical Studies of Biomolecules at the NIH

National Institute Of Diabetes And Digestive And Kidney Diseases

Investigators

Linked publications & trials

Abstract

The Covid-19 pandemic proved quite disruptive to our core research program, and led to collaborative studies that focused on mechanism of disease transmission via aerosols, such as SARS-CoV-2. Our pioneering studies with Dr. Ad Bax to visualize speech droplets via laser scattering and evaluate their potential role in the spread of the SARS-CoV-2 virus led to another collaboration with Dr. Christopher Koh. First, a bit of background. The Anfinrud group operates an ultrafast time-resolved laser spectroscopy lab in Bldg. 5, Rm. B2-10, which is outfitted with HEPA filtered airflow over the laser tables to keep the optics clean. One section of the laser table that has traditionally been reserved for developing new experimental approaches was cleared and a laser scattering apparatus capable of visualizing speech droplets emitted from one's oral cavity was set up. Our initial studies employed a multi-watt CW Nd:YLF green laser whose output was directed through custom optics designed to generate a thin vertical light sheet that was routed along the length of an optical table. The light sheet passed through slits cut into the sides of a cardboard box whose interior was painted black. When particles passed through the light sheet, bright flashes of light appeared and were recorded with an iPhone camera. Speaking into the light sheet with the room lights shut off revealed a surprisingly large number of speech droplets that lingered in the air for many minutes. These results were published in a paper entitled "Visualizing Speech-Generated Oral Fluid Droplets with Laser Light Scattering" in the New England Journal of Medicine, April 15 (2020), and another entitled "The airborne lifetime of small speech droplets and their potential importance in SARS-CoV-2 transmission" in the Proceedings of the National Academy of Sciences, USA, 117, 11875-11877 (2020). We subsequently fought off a challenge to the validity of our work in a publication entitled "SARS-CoV-2 transmission via speech-generated respiratory droplets," which was published in the Lancet Infectious Diseases, Sept. 11 (2020). This work led to an invitation to submit a review article which we entitled "Breathing, speaking, coughing or sneezing: What drives transmission of SARS-CoV-2?" which was published in the Journal of Internal Medicine, 08 June (2021). A follow-up publication entitled "Hybrid measurement of respiratory aerosol reveals a dominant coarse fraction resulting from speech that remains airborne for minutes" was published in the Proceedings of the National Academy of Sciences USA 119, e2203086119 (2022). Having learned of our studies, Dr. Christopher Koh, acting director of Clinical Research, NIDDK, approached us to explore the possibility of using our light scattering methods to visualize aerosolized particles generated by endoscopy procedures. To pursue this study, we developed a laser scattering apparatus based on a very compact and cheap solid-state laser diode that generates blue light, which scatters more efficiently than green light. The diode was purchased as a discrete electronic component that had to be packaged, powered, and integrated into an optical system that created a light sheet with dimensions appropriate for the task. A novel packaging design was conceived that provides for efficient cooling of the diode when operating with multi-watt power output. A horizontal light sheet was directed across an optical table and near a structure that supported a lamb esophagus, with cameras mounted to visualize particles that pass through the light sheet. Flashes of light were observed when inserting and retracting endoscopic tools through a septum on the endoscope, and when inserting and retracting the endoscope into the lamb esophagus. The aim was to evaluate which, if any of these procedures was capable of generating aerosols that, if the patient was infectious, could lead to transmission of disease. The results of this study entitled "Visualizing Endoscopy- Generated Aerosols with Laser Light Scattering" was recently accepted for publication in Gastrointest. Endosc. (2022). To better understand airborne transmission of disease, we need to further our understanding of mechanisms for generating droplets from potentially-infectious bodily fluids, their emission pathways, the size cutoff for becoming aerosolized due to evaporation of water, the length of time these aerosol particles linger in the air, and the probability they can be inhaled by an unsuspecting individual in the same vicinity, whether outdoors or indoors. We have developed a new particle scattering instrument that should prove capable of quantitatively characterizing the size distribution of droplets emitted from the oral cavity and their airborne lifetime. The basic idea is to introduce droplets into a humidity-controlled, vertically-oriented tube and record the time it takes them to fall and pass through a light sheet near the bottom of the tube. The time of flight is inversely proportional to the square of the size of the particle, as is the transit time required to pass through a light sheet of known thickness. Heating the top and cooling the bottom of the tube produces a stable, precisely defined temperature gradient that ensures particles dont get caught up in updrafts that could arise from room temperature fluctuations. The air in the tube is analogous to the medium inside a chromatography column: the largest particles fall through the light sheet first, and the smallest particles fall through last. The size of particles that can be characterized with this design spans a much larger range than is covered by commercially-available particle sizing instruments. This instrument has been built and preliminary studies suggest it should be capable of achieving many of the objectives that led to its design. However, the post-doc working on this project left to take a position with the FDA, and that project is currently on hold. As the NIH began loosening requirements for returning to the lab, the pace of our collaborative pressure-jump NMR studies has picked up. Three home-built pressure-jump systems are currently operable. The recent upgrade of consoles used to operate the high-field Bruker NMR magnets required some modifications to the pressure-jump apparatus control systems, and the software that operates them has also been tweaked to address some other issues that came to light during their operation. Dr. Baber, a staff scientist who supports NMR efforts in LCP is currently being trained to take over day-to-day oversight of these systems and has proven a very capable and welcome addition to this collaboration. Together, we have improved and standardized the design of the high-speed pneumatic valve controllers used to rapidly switch the sample cell pressure. Moreover, he assembled additional transfer lines so we always have a spare ready to go at each installation. Now, when a leak develops during an experiment, it is relatively straightforward to swap out the transfer line and resume data acquisition with minimal disruption. We aim to update the computer control software currently used to operate the pressure-jump apparatus and implement a more modular, hierarchical, and extensible structure along the lines we are developing to operate the BioCARS beamline at the APS. However, that transition will await the time when our improved approaches have been fully commissioned at that site.