Detoxicated Vaccine.—Another variety of vaccine, in which the bacterial protoplasm has been so modified as to remove its toxicity, as well as its infectivity, is known as "detoxicated." Pasteur's anthrax cultures incubated at an unsuitable temper ature, and Raw's tubercle cultures grown for a long period of years upon artificial culture media, were detoxicated in this sense by purely biological methods. Recent workers have attempted to effect the same object by treating the bacterial protoplasm with either mineral acids or alkalis. These chemical methods, how ever, profoundly change the protoplasm of the organism, and the injection of the resulting vaccines into the animal economy is not as a rule followed by the production of any large amount of the "specific" antibodies as at present recognized. That detoxi cated vaccines of this type do subserve a useful purpose is beyond question, however, although the mechanism which is here brought into action is probably closely allied to that of protein shock (see IMMUNITY).
Mode of Preparation.—All the above varieties of vaccines are prepared on very similar lines. The selected organism is culti vated usually on a solid substratum, under optimum conditions for that particular period which will give the maximum develop ment, and the resulting growth is emulsified in normal saline solution. In the preparation of sensitized vaccines the appro priate anti-serum is added to the emulsion and the mixture is allowed to stand at o° C for a period of about 12 hours. Next, the amount of bacterial protoplasm held in suspension per unit volume (I c.c.) is measured, and several methods are avail able for the purpose. Undoubtedly the most accurate method involves drying and weighing the mass of bacteria, but this is a lengthy process, and often the time factor is of paramount im portance where therapeutic vaccines are concerned. For this reason the plating methods ordinarily employed for the estimation of the number of bacteria suspended in fluids, such as water, are rarely resorted to, and other more rapid methods have been utilized. Thus Sir Almroth Wright mixed equal volumes of the bacterial emulsion and blood from a normal individual, and by means of microscopical films estimated the ratio existing between the red cells and the bodies of the bacteria in the mixture. This ratio having been ascertained, a simple calculation determines the number of bacteria in the unit volume, since the number of erythrocytes normally present in man is a remarkably constant factor (amounting to 5,000 million per cubic millimetre). Again, the bacteria may be counted by means of the ordinary haemo cytometer, or the estimation may be made by comparing the suspension of bacteria with standard tubes containing opaque fluids corresponding to definite numbers of various organisms per cubic centimetre.
In most instances the microbes contained in the emulsion are now altered by heat coagulation (by suspension in a water bath adjusted to the thermal death point of the particular organism) by autolysis, or by lysis due to the action of chemicals such as sodium fluoride or carbolic acid, so that their infectivity is de stroyed; in the case of sensitized vaccines the microbes may, at the discretion of the bacteriologist, be left in the living state and unaltered—save for their combination with the corresponding antibody. In the chemically detoxicated vaccines the solution of the bacterial protoplasm is effected at this stage by the ad dition of the requisite acid or alkali, and the vegetable protein is further purified by repeated precipitation and solution.
Finally, the emulsion, is adjusted to some pre-determined standard by dilution or concentration, a small quantity of a preservative, such as tricresol or phenol, is added and the finished product stored in suitable receptacles.