The unstable ice nucleation properties of Snomax® bacterial particles

Abstract

Snomax® is often used as a surrogate for biological ice nucleating particles (INPs) and has recently been proposed as an INP standard for evaluating ice nucleation methods. We have found the immersion freezing properties of Snomax particles to be substantially unstable, observing a loss of ice nucleation ability over months of repeated droplet freezing measurements of the same batch of Snomax stored as dry pellets in a freezer. This reflects the fragility of the most ice active large protein aggregates and presents issues for the use of Snomax as an INP standard. The ice nucleation properties we determined using a fresh Snomax batch agreed well with the recent method intercomparison from the Ice Nucleation UnIT (UNIT) project, while an older batch did not. Using an oil immersion droplet freezing technique, repeated freezes of Snomax droplets resulted in a decrease in ice nucleation ability after successive refreezes. We attribute this to the disruption or displacement of the most ice active protein aggregates that are thought to contain the ice nucleants. Partitioning of the protein aggregates from the droplet into the immersion oil that is accelerated by droplet freezing events could explain the observed decrease in freezing ability. Droplets in mineral oil or low viscosity silicone oil experienced a smaller reduction in freezing temperature than when squalene oil was used. The effect of the immersion oil may be specific to proteinaceous biological particles, and we have not observed it in nonproteinaceous materials. Caution is warranted in the use of oil immersion droplet freezing methods to determine immersion freezing properties.

Effect of particle surface area on ice active site densities retrieved from droplet freezing spectra

“…A notable difference from the droplets containing illite is that there is significant weakening in ice nucleation ability as the concentration/surface area of Snomax is reduced. This behavior matches what is known regarding the low abundance of the most efficient but fragile Type I ice nucleating proteins that freeze at −3 to −2 • C vs. the more abundant and resilient but less efficient Type III proteins that freeze around −8 to −7 • C (Polen et al, 2016;Turner et al, 1990;Yankofsky et al, 1981).…”

“…Figure 6a shows decreasing concentration freezing curves for droplets containing Snomax particles. Snomax is a freeze-dried powder manufactured from nonviable Pseudomonas syringae bacteria and is commonly used to make artificial snow due to its very mild freezing temperature of −3 to −7 • C. Its ice nucleation properties are attributed to large protein aggregates, and Snomax is often used as a proxy for atmospheric biological INP (Pandey et al, 2016;Polen et al, 2016;Wex et al, 2015). A similar approach was undertaken in which g was retrieved using the highest concentration freezing curve (solid blue line).…”

“…We investigated a similar trend when freezing droplets containing commercial Snomax (York International), and MCC cellulose (Sigma-Aldrich) particles immersed in oil in our in-house cold plate system, described by Polen et al (2016). The relevant system details are that particlecontaining water droplets of approximately 500-700 µm in diameter are immersed in squalene oil, analogous to the method of , and the droplets' freezing temperature is determined optically during a constant 1 K min −1 cooling cycle.…”

Leipzig Ice Nucleation chamber Comparison (LINC): intercomparison of four online ice nucleation counters

“…The birchS sample seems to lose ice activity during storage, suggesting that care should be taken when comparing results of ice nucleation of biological samples because some samples could potentially change over time. A loss of ice activity over time has also been previously observed for Pseudomonas syringae (e.g., Polen et al, 2016).…”

A new multicomponent heterogeneous ice nucleation model and its application to Snomax bacterial particles and a Snomax–illite mineral particle mixture

“…Biological particles such as Pseudomonas syringae bacteria can induce freezing in supercooled water at temperatures as high as −2 • C Polen et al, 2016;Wex et al, 2015). While mineral dust particles are the leading candidate for the most abundant ice nucleating particles (INPs) in the atmosphere (Hoose et al, 2008;, lidar and radar measurements (Bühl et al, 2013), as well as laboratory studies in which melted precipitation samples were refrozen (Christner et al, 2008;Petters and Wright, 2015;Vali, 1971Vali, , 1996, have shown that freezing can occur at temperatures higher than −12 • C. In this temperature range, no investigated mineral samples of atmospherically relevant particle sizes (or other atmospherically relevant non-biological particles) have exhibited detectable ice nucleating activity in Atkinson et al (2013), Cantrell and Heymsfield (2005), DeMott et al (2015), and Murray et al (2012).…”