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Endangered species that have never before been able to be preserved could soon be protected. Scientists have solved the mechanism behind the long-term freeze storage technique ‘cryopreservation’ that is used to preserve the embryos of species that are in danger of extinction. The results are published in the 2 March 2016 edition of ‘Royal Society Open Science’.

Despite global conservation efforts, many marine, fresh-water and land animals may become extinct in the next few decades. In an effort to protect endangered species, conservationists can freeze embryos in the early stage of their development and these can be stored for thousands of years - a technique known as cryopreservation. Some species’ embryos, however, are more difficult to cryopreserve than others. For the process to be successful, it is essential that ice crystals do not form as they act as tiny knives, puncturing cell membranes and killing the embryo.

Now for the first time, the mechanism behind cryopreservation has been cracked at the molecular level. These results will make it possible to cryopreserve embryos of endangered species which are currently unachievable. In mammals the freezing of embryos is common place, whereas in some fish species it is more difficult; fish eggs have strong outer membranes that are impermeable to cryoprotectants - the chemicals used to preserve the embryo and protect it from ice crystals.

In neutron experiments at STFC’s ISIS Neutron and Muon Source, scientists ‘watched’ the freezing process take place and were able to see what was happening to individual components in the mix of cryoprotectants and water.

They found that in mixtures with a particular concentration of cryoprotectants, the chemicals formed long chains and acted like a mesh or ‘sponge’ that locked the smaller water molecules in to pockets. In normal freezing, water molecules link up to form ice crystals, however the cryoprotectant mesh meant the water molecules were isolated in small clusters.

The study confirms observational experiments by scientists in the Czech Republic, who after several years of trial and error experiments, noticed that a particular mixture of cryoprotectants led to a perfectly preserved fish egg from a Common Carp.

Now the mechanism of cryopreservation has been found, these results can go towards a model for the cryopreservation of endangered species with more challenging embryos, such as some coral fishes.

Professor William Holt, former Head of Reproductive Biology at the Zoological Society of London (ZSL) said this work will contribute to conservation efforts:

"This represents a significant step in the development of a technique for fish oocyte cryopreservation. If eggs could be frozen and stored without losing viability, they would provide a valuable resource for conservation programmes. By providing genetic support to endangered populations, these techniques would help reduce the damaging effects of inbreeding, enhancing animal welfare."

The experiments took place on ISIS instrument, SANDALS. In the experiments, Dr. Alan Soper and co-author Dr. Oleg Kirichek took three different mixtures of cryoprotectants and quenched cooled, or rapidly cooled them in liquid nitrogen to -196 degrees celsius in seconds. They then performed neutron diffraction experiments to see the structure of each mixture after freezing.

As neutrons are very sensitive to hydrogen, a key element of water, scientists were able to ‘see’ at the molecular level what happened to the mixture after quench cooling.

Dr. Alan Soper, an ISIS scientist and world expert in the structure of water and water-based solutions who was co-author on the study said:

"Neutrons were crucial for this study because they instantly highlighted any ice that did develop in some samples, and also, by virtue of the ability to perform hydrogen/deuterium substitution on the water molecules, allowed us to access the water structure independently of the contributions from the other components in these solutions.”

STFC supported the neutron scattering part of the research, both through funding beam time on ISIS and through the provision of samples.

For full details see the paper.

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ISIS

ISIS is a world-leading centre for research in the physical and life sciences at the STFC Rutherford Appleton Laboratory near Oxford in the United Kingdom. Our suite of neutron and muon instruments gives unique insights into the properties of materials on the atomic scale. We support a national and international community of more than 3000 scientists for research into subjects ranging from clean energy and the environment, pharmaceuticals and health care, through to nanotechnology and materials engineering, catalysis and polymers, and on to fundamental studies of materials.

We use the technique of neutron scattering. Neutrons tell us where atoms are and how they are moving. By studying how materials work at the atomic level, we can better understand their every-day properties – and so make new materials tailor-made for particular uses. ISIS also produces muons for use in a similar way, providing additional information on how materials work at the atomic scale.

The Zoological Society of London (ZSL)

Founded in 1826, the Zoological Society of London (ZSL) is an international scientific, conservation and educational charity whose mission is to promote and achieve the worldwide conservation of animals and their habitats. Our mission is realised through our ground-breaking science, our active conservation projects in more than 50 countries and our two Zoos, ZSL London Zoo and ZSL Whipsnade Zoo.