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Isolation of a Vaccinia-specific Hapten from Vaccinia-infected Sheep Dermis

Department of Virology, St Mary's Hospital Medical School, London, W. 2

C. J. M. Rondle

Department of Bacteriology and Immunology, London School of Hygiene and Tropical Medicine, London, W.C. 1

Virus-free buffer extracts of vaccinia-infected sheep dermis were fractionated by elution from DEAE-cellulose with increasing concentrations of NaCl. The fraction eluted with 4.0M-NaCl was serologically homogeneous when examined in precipitation-in-gel tests. Further fractionation on Sephadex G-200 gave a material which was also physically homogeneous when examined by both ultracentrifugal and electrophoretic techniques. A sedimentation coefficient of 3.47 S and a molecular weight of 31,000 were calculated from ultracentrifugal data. The isolated material contained 74.3% DNA, 9.4% protein estimated as bovine serum albumin and 14.9% carbohydrate estimated as glucose. The DNA contained the bases adenine, thymine, guanine and cytosine in the ratios 1.0:1.1:0.8:0.8. Paper chromatography of formic-acid hydrolysates of the material resolved seven substances reacting with ninhydrin. Qualitative colorimetric tests indicated, apart from deoxyribose, the presence of a hexose and a hexuronic acid.

The serological activity of the isolated material was heat-stable and identical with that of a previously described heat-stable extract of vaccinia-infected rabbit dermis. Treatment with enzymes indicated the presence of two different, serologically active sites. Failure to elicit an antibody response following injection into rabbits suggested that the isolated material was a hapten. Serum absorption studies showed serological identity with an antigen present on the surface of the vaccinia virus particle.

Received 22 March 1968; accepted 13 May 1968.