A method for separating and quantifying organic and inorganic ice nucleating substances in atmospheric samples based on density gradient centrifugation

Ice nucleating substances (INSs) influence the properties and frequencies of ice and mixed-phase clouds in the atmosphere, and hence, climate and the hydrological cycle. INSs can be classified as inorganic (e.g., mineral dust, volcanic ash) or organic (e.g., bacterial cells, cell-free proteins). While the properties of both INS classes have been studied in the laboratory, the amounts in the atmosphere are still poorly constrained. Here, we demonstrate a new method for separating and quantifying inorganic and organic INSs. First, INS suspensions were separated into a high-density isolate containing inorganic INSs and a low-density isolate containing organic INSs using density gradient centrifugation, and then INSs were quantified in each isolate using a droplet freezing technique. Inorganic K-feldspar and organic Snomax INSs were used to test our method. The average K-feldspar INS recovery in the high-density isolate was 54%, with no evidence of K-feldspar INSs in the low-density isolate. The average Snomax INS recovery in the low-density isolate was 27%, with small amounts of Snomax contaminating the high-density isolate. A mixture of K-feldspar and Snomax was successfully separated, with recoveries comparable to those observed for K-feldspar and Snomax individually. Recoveries less than 100% can be explained by losses of INSs to vessel walls, accidental mixing of the different density layers during pipetting, and incomplete collection of material during pipetting.