Field-compatible tissue cryopreservation enables delayed endothelial cell culture

Decades of biomedical research have developed robust tools in molecular and cell biology, including cell culture systems, for model vertebrate species. Logistical constraints associated with remote fieldwork limit the application of these techniques to non-model species. Traditionally, cell culture requires tissue collection from live or very recently deceased animals, followed by immediate sterile processing and cell isolation, but these requirements are often impractical in field settings. To address this limitation, we developed a field-compatible method to cryopreserve arterial tissue from pinnipeds for delayed endothelial cell isolation. Grey seal placentae were opportunistically collected, and arteries were preserved using benchtop blunt dissection and a portable ​​-80 °C freezer with a standard cell freezing medium. Samples were held in a portable ​-80 °C freezer for ~ 2 months before transfer to LN 2 vapor storage for ~ 1 month, followed by endothelial cell isolation. Cells isolated from cryopreserved arteries are phenotypically endothelial, proliferate in culture and are the first endothelial cell cultures derived from grey seals. Grey seal endothelial cells respire after culture expansion, and ATP production rates are ~ 1.5×​​ greater than those in freshly derived endothelial cell cultures from a related species, though the relative contributions of OXPHOS and glycolysis were comparable between cell preparations. This method is practical for field researchers and supports the generation of primary cell cultures weeks-to-months after initial tissue collection with minimal resources, strengthening the ability to investigate the cellular basis of physiological phenomena in the natural world.