Preparation of SEM hydrogel samples using a high pressure water freeze fracture method

ABSTRACT The preparation of SEM samples is a complex and time-consuming process. The present report describes the development of a simple and inexpensive high pressure water freeze fracture method for the preparation of SEM samples from hydrogels without using organic solvents. This method involves two key features. First, the sample and water are frozen with liquid nitrogen in a pressure-resistant container, and the sample is pressurized by volume expansion due to phase transition of the water around the sample as it freezes, which suppresses the growth of ice crystals due to the increase in viscosity of the water inside and around the sample. Second, a cross-section of the sample is prepared without the use of a knife. This method is expected to contribute significantly to biological and food science research.