We have shown that it is possible to obtain from extracts of vaccine virus-infected tissues a substance or substances, apparently protein, which are serologically active, and which are specifically related to vaccinal infection. The present investigation is concerned with a study of their immunological reactions. Their intraperitoneal injection in rabbits is followed by the appearance in the serum of these animals of antibodies directed specifically against them. The precipitating capacities of this serum are entirely removed after addition of appropriate quantities of heated vaccine virus extract, indicating that antibodies against only the heat-stable antigens have been produced. Further evidence of the specificity of the antibodies is gained from the reverse experiment, that is, absorption of a virus extract with the serum. We have shown that under suitable conditions the serum will remove S antigen specifically leaving the labile substances in solution. This would apparently indicate that they are serologically distinct although both are vaccinal products.

Serum of animals injected with the S substance and containing antibodies against it in high titer is apparently capable of neutralizing minute amounts of active virus. The animals providing the serum are, however, without demonstrable resistance to vaccinia. The significance of the neutralizing activity of the serum is debatable because it is of a greatly different order of magnitude from that which follows infection with vaccinia.

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