Babesia major in Britain: infectivity for cattle of cryopreserved parasites derived from Haemaphysalis punctata nymphs.

Babesia major-infected Haemaphysalis punctata nymphs, prefed for four days on rabbits, were ground in a modified Eagle's minimum essential medium containing N-2 hydroxyethylpiperazine-N1-ethane-sulphonic acid (HEPES) buffer using sterile sand or ground glass as triturating agents. The resultant supernatant fluids were shown to be infective for splenectomised calves. When attempts were made to cryopreserve the fluids using either dimethyl sulphoxide or glycerol to a final concentration of 10 per cent, only the former successfully maintained the intravenous infectivity of the organisms released by grinding with glass. A cooling rate of approximately 1 degrees C/min resulted in successful cryopreservation but a more rapid cooling rate did not.