In 1972, David Whittingham, Stanley Leibo, and Peter Mazur published the paper, “Survival of Mouse Embryos Frozen to -196 ° and -269 °C,” hereafter, “Survival of Mouse Embryos,” in the journal Science. The study marked one of the first times that researchers had successfully cryopreserved, or preserved and stored by freezing, a mammalian embryo and later transferred that embryo to a live mouse who gave birth to viable offspring. Previously, scientists had only been successful cryopreserving single cells, like red blood cells. Mammalian embryos, on the other hand, were more difficult to cryopreserve because they are more complex and therefore more easily weakened or destroyed by the formation of ice within its cells. Whittingham, Leibo, and Mazur’s work provided a successful model for mammalian embryo cryopreservation, a technology that later expanded to cryopreserve more complex embryos, such as human embryos.

Peter Mazur was a researcher in the US who developed new ways of preserving biological material by freezing it, a process called cryopreservation. If done correctly, cryopreservation enables scientists to store or study biological material for an extended period of time. If done incorrectly, cryopreservation can easily harm or destroy biological material. Mazur worked to find the best ways to cryopreserve different cells, embryos, and organs in order to minimize the damage caused by freezing. Throughout the 1960s and 1970s, Mazur and his colleagues published a series of papers that ultimately led to the discovery of previously unexplored factors that can cause harm to cells during the cryopreservation process. He called that discovery the two-factor hypothesis. That same year, Mazur also contributed to one of the first successful attempts at cryopreserving viable mouse embryos. Mazur’s work to improve the cryopreservation process helped to establish foundational knowledge that was later used in many different fields, such as reproductive health and conservation.

Stanley Paul Leibo studied the cryopreservation of embryos in the US in the twentieth century. Cryopreservation is a method of preserving biological material through freezing. Early in his career, Leibo collaborated with other scientists to study why cells were oftentimes injured during freezing. Later, Leibo and his team accomplished one of the first successful births using previously-frozen mammalian embryos. Leibo continued evolving simpler and more reliable methods of cryopreservation and embryo transfer for many different species over the course of his career, such as the development of a one-step procedure of transferring fertilized embryos between cattle. Leibo’s work to develop simple and reliable ways to cryopreserve cells and embryos enabled its use in a wider scope of research, including agriculture, reproductive medicine, and conservation.

In 1972, Peter Mazur, Stanley Leibo, and Ernest Chu published, “A Two-Factor Hypothesis of Freezing Injury: Evidence from Chinese Hamster Tissue-culture Cells,” hereafter, “A Two-Factor Hypothesis of Freezing Injury,” in the journal, Experimental Cell Research. In the article, the authors uncover that exposure to high salt concentrations and the formation of ice crystals within cells are two factors that can harm cells during cryopreservation. Cryopreservation is the freezing of cells to preserve them for storage, study, or later use. Mazur originally suggested the two factors in a 1970 paper, but that article was based on evidence from simple yeast cells. By using hamster cells in 1972, Mazur, Leibo, and Chu confirmed that Mazur’s two-factor hypothesis applied to more complex mammalian cells. The article dispelled the widely accepted notion that rapid cooling rates were safest for all cells, and instead showed that each kind of cell had a different optimal cooling rate depending on the solution in which it froze.

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