An Experimental Technique for Accurate Measurement of the Freezing Point of Solutions and Ice Melting in the Presence of Biological Objects on the Examples of P. Syringae and E. Coli

Pseudomonas syringae is a widely distributed bacterial epiphyte of plants. When the temperature of the environment drops below zero, P. syringae can serve as biological ice nuclei due to the presence of specialized ice-nucleating proteins. This property has found application in various fields, but despite its evident importance, the molecular mechanisms behind protein-induced freezing have remained largely elusive. One of the problems in the study of the ice-nucleating process is the difficulty of carrying out experiments with freezing/melting solutions at near-zero temperatures. The experimental design implies special requirements for the equipment used and measurement technique. In this work, we used an experimental setup assembled from a dry thermostat that maintains a constant temperature and an accurate digital thermometer. We experimentally tested the possible sources of errors of the setup. As a result, we have shown that the accuracy of determining the freezing temperature of liquids and the coexistence of ice and water is mainly determined by the accuracy of the thermometer. The accuracy of determining the melting point of ice depends on the volume of the sample and is systematically underestimated in our setup. Using the proposed experimental technique, we performed a comparative study of P. syringae and E. coli cells, which revealed that P. syringae cells affect not only the freezing point of the solution but also the temperature of the coexistence of ice and water. The observed effect can be explained by the binding of P. syringae cells to the ice surface.