Cryogenic Light Microscopy and the Development of Long-term Cryopreservation Techniques for Fungi

Since Henry Power in 1663 successfully revived eelworms which had been frozen in vinegar for a few hours, there has been much interest in cryopreservation. As liquid nitrogen and other cooling agents became more widely available a standard method involving a single cooling rate for the cryopreservation of fungi was proposed by Hwang in 1960. This method was used until the early 1980s, but an increasing number of recalcitrant strains were found. Research at the time was empirical and based on attempts to find the universal cryoprotectant which would protect all fungi irrespective of the cooling rate. The real need was to go back to basics and study the response of the living cell to freezing and thawing. The use of cryogenic light microscopy to study fungi provided a major breakthrough, and showed that cooling rates which cause shrinkage or ice formation should be avoided. Further work is required particularly on membrane structure and the mechanism by which intracellular ice causes injury, as well as the effects of the age of the culture, hyphal structure and growth requirements. Cryopreservation is the only method of preservation which will store fungi and other cells for extremely long periods of time without detectable genetic or morphological change.