Proceedings: Preservation and storage of microorganisms in the gas phase of liquid nitrogen

Methods are described which permit the rapid preparation and retrieval of suspensions of microorganisms stored in the gas space of a liquid nitrogen refrigerator.Inocula are currently used for assay plates, turbidimetric assays, bioautograph plates, and various fermentation processes. The most suitable suspending agents, 10% glycerol with 5% of either lactose, maltose, or raffinose, were devised by determining viabilities from alternate freezing and thawing of cell suspensions.Four milliliters each of concentrated cell suspensions were pipetted into cotton-plugged or plastic-capped glass tubes (13 × 100 mm). After rapid noncontrolled freezing in the nitrogen gas phase, the tubes were stored horizontally either in test-tube racks or in bulk containers.Frozen suspensions of vegetative cells of some algae, many genera of bacteria, as well as spores, vegetative cells, and fragmented mycelia of streptomycetes and fungi have been conveniently prepared from either agar or broth cultures. Sufficient inocula have been prepared to be used for periods of a few weeks to several years.