Although the presence of antifreeze and ice nucleating agents in the hemolymph of insects has been well documented, there have been no reports of either of these types of agent in vertebrates. The technique of differential scanning calorimetry was used to examine the blood, serum, and plasma of a freeze-tolerant frog, Rana sylvatica, for the presence of antifreeze protein activity. Results demonstrate the absence of antifreeze protein but the presence of an ice nucleating agent that may serve as a functional component of the overwintering strategy of this species. Ice nucleating activity was detected in samples of cell-free blood, serum, and plasma, suggesting that the agent is a soluble component and possibly plasma protein. To our knowledge, the identification of ice nucleating activity in this freeze-tolerant vertebrate is novel.

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Journal Cryobiology
Wolanczyk, J.P. (Jan P.), Storey, K, & Baust, J.G. (John G.). (1990). Ice nucleating activity in the blood of the freeze-tolerant frog, Rana sylvatica. Cryobiology, 27(3), 328–335. doi:10.1016/0011-2240(90)90032-Y