The known host range of naturally-occurring transmissible spongiform encephalopathies has expanded in recent years to include wild ruminants. Chronic wasting disease (CWD) occurs in mule deer (Odocoileus hemionus hemionus) and Rocky Mountain elk (Cervus elaphus nelsoni) in Colorado and Wyoming, United States of America. These species belong to the family Cervidae. Cases have occurred primarily in captive animals but a few affected free-ranging animals have been identified. Clinical disease in both species is characterised by progressive weight loss, behavioural alterations and excessive salivation. In deer polydipsia and polyuria also commonly occur. Significant lesions are confined to the central nervous system and consist of spongiform change in grey matter, intraneuronal vacuolation, astrocytosis and amyloid plaques. Inflammatory reaction is absent.The origin of this disease is not known. In contrast to the cases of spongiform encephalopathy recognised in five species of antelope (family Bovidae) in British zoological parks, which are an extension of the current bovine spongiform encephalopathy epizootic, CWD is not the result of food-borne exposure to the infectious agent. CWD appears to be maintained within captive populations by lateral and, possibly, maternal transmission.Spongiform encephalopathies in wild ruminants are currently geographically isolated and involve relatively small numbers of animals. However, these potentially transmissible diseases could be of greater importance in the future and should be viewed with concern in the light of international movements of wild ruminants and the current expansion of the game farming and ranching industry in many parts of the world.

Most attention to the transmissible spongiform encephalopathies (TSEs) in animals has been directed at the domestic animal diseases scrapie and bovine spongiform encephalopathy (BSE). This is due to the economic value of the animals, and in the case of BSE, because of the causal relationship between BSE and the human disease, variant Creutzfeldt-Jakob disease (vCJD). Transmissible spongiform encephalopathies also occur in non-domestic animals; one of these, transmissible mink encephalopathy (TME), was the second animal TSE recognised following scrapie, the oldest known TSE. In each of these diseases, oral exposure to the agent, in various forms, constitutes the most significant route of infection.

A modified centrifugal élutriation technique is described for the isolation of large num bers of lymphocytes and monocytes. Elutriation was carried out by lowering the rotor speed at a constant flow rate which was generated by hydrostatic pressure. The flow rate could be kept constant if the separation procedure was performed at high pressure and high systemic resistance.Up to 2.3 X 109 mononuclear cells derived from 2000 ml blood were separated in one single experiment in approximately 1 h. The lymphocytes and monocytes were isolated at purities of 98 ± 1% and 94 ± 1% respectively. The purity of the lymphocytes was increased to 99.8 ± 0.1% by a second élutriation run. Additional advantages of the élutriation procedures are that the choice o f medium is free, and that relatively large numbers of cells may be separated with high recoveries.

One of the major disadvantages of centrifugal elutriation (CE) is the relatively large volume (150 ml) of the various fractions, especially if small numbers of cells have to be separated and the fractions contain few cells.To reduce the volume of the fractions 2 elutriator rotors were coupled in series. Since the rotor speed of the second rotor was always kept 750 rpm liigher than that of the first rotor, cells elutriated from rotor 1 were collected in rotor 2. After elutriation of a complete fraction from rotor 1, and collection in rotor 2, the cells were harvested from rotor 2, This was achieved by means of a flow distribution unit (FDU), which made it possible to disconnect the flow of both rotors and simultaneously reverse the flow of the second rotor.It is demonstrated that 40-95 X lO 6 mononuclear leukocytes may be fractionated without loss of resolution in fractions of only 9 ml. The lymphocyte ( > 99%) and monocyte subpopulations (88-94%) obtained were as pure as with CE carried out with only 1 rotor. In addition, the cells in rotor 2 could be washed and suspended in culture medium prior to harvesting by means of the FDU. In this way loss of cells by additional centrifugation steps was avoided, Erythrocytes (RBC) present in certain lymphocyte fractions were lysed with N H 4C1 and after lysis of the RBC and elution of ghosts and debris, the cells were washed and harvested. This procedure did not affect cell viability and the PHA response of the lymphocytes.The versatile system described made it possible to apply CE for the separation of small numbers of cells without loss of resolution, and demonstrated that CE is ideally suitable for concentration and washing of cells, and removal of contaminating RBC, not affecting the recovery, viability and function of the cells.

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