title: Successful cryopreservation of adult male Onchocerca gutturosa in liquid nitrogen.
abstract: Experiments involved the use of methanol and ethanediol (ED) as potential cryoprotectants for the storage of adult male Onchocerca gutturosa at low temperatures. Worms were incubated with or without 20% (v/v) methanol for 15 min at 0 degrees C before cooling to -10 degrees, -25 degrees or -79 degrees C. Results of trials of storage from 1 to 14 days showed that prolonged survival could only be obtained by storing at -79 degrees C without the addition of cryoprotectant. However, worms were also successfully cryopreserved in liquid nitrogen (-196 degrees C) using a simple 2-step procedure in which ED was used as an essential cryoprotectant. This schedule involved an initial incubation of parasites in serum free media with 10% (v/v) ED at 37 degrees C for 15 min, followed by incubation in 40% (v/v) ED for 10 min at 0 degrees C on crushed ice. Worms were then transferred individually into 20 microliter drops of 40% (v/v) ethanediol on glass coverslips at 0 degrees C and immediately plunged into liquid nitrogen. A rapid thaw of the parasites resulted in normal motility and survival levels equivalent to unfrozen controls. This technique will be useful for the long-term storage and transportation of this and possibly other species of Onchocerca parasites.
mesh_terms: D000818:Animals; D003451:Cryoprotective Agents; D005026:Ethylene Glycols; D005615:Freezing; D008297:Male; D000432:Methanol; D009584:Nitrogen; D009854:Onchocerca; D011309:Preservation, Biological
chemical_list: D003451:Cryoprotective Agents; D005026:Ethylene Glycols; D009584:Nitrogen; D000432:Methanol