Various modifications of this procedure have been made. To avoid the inevitable entrance of ærial germs, flamed pipettes have been inserted into a puncture in the turgid lung, made with a flamed knife, the point is then broken and the serum is drawn up to fill the vacuum caused by the heat employed in sterilization. The point is then sealed by melting the glass in the flame of an alcohol lamp, or by melted wax.
Nocard washed the surface of the infiltrated or hepatized lung with boiled water, then with a sterilized knife cut out a deep segment so as to leave a conical cavity and covered this with an awning to keep off dust. The space soon fills with the draining exudate in a very pure condition.
Pasteur inoculated a calf in the dewlap or behind the elbow, where an enormous engorgement forms, involving the whole ventral aspect, and supplying an almost unlimited amount of serum, which may be collected with such precautions against contamination as are indicated above. The product is thus secured at a comparatively cheap rate, and the risk of its diffusion on the air in breathing is lessened enormously.
Another method is to employ the serum from the swellings in the inoculated tails and carry it on indefinitely from tail to tail.
Arloing in his turn employed the cultures in vitro of his pneumo-bacillus, but with the modern evidence that this is not the infective germ, such cultures can not always be implicitly relied on.
In place of the hypodermic syringe, Australians have used a simple thread soaked in the exudate and drawn through beneath the skin. Others have used a small lancet with a groove hollowed out in the middle of the flat surface of the blade on one side, and which carries in the required drop of the serum.
To preserve the exudate for some time against decomposition, it has been kept on ice, or mixed with chloral hydrate or phenic acid (½ its volume), and glycerine (½ its volume) for two or three months.
Injection into the veins practiced by Burdon-Sanderson has the advantage of producing no local lesion whatever and yet securing a fair measure of immunity. It is, however, a much more delicate operation as it entails a possible though remote possibility of producing capillary thrombi, and some danger of infection of the wound in the vein. To avoid this latter, the jugular vein is raised as for phlebotomy, and a short needle is passed into it. A longer and more delicate needle is now passed through this, and the injection of a few drops of the exudate is made through the latter. The small needle is then carefully withdrawn to be followed a few seconds later by the large.
Injection of the Sterilized Exudate Subcutem. In 1881 having found that liquids obtained from old sequestra, produced no local lesions when inoculated subcutem, but secured immunity for the subjects, I inoculated ten susceptible cattle with the fresh pulmonary exudate which had just been heated for 30 minutes to 180° F. No local inflammation nor exudate occurred in any of them, but subsequent inoculations with fresh unsterilized lung plague exudate were resisted in the same way. Six of these immunized cattle were subsequently placed in two infected stables and herds, (Mr. Butts, Brooklyn, N. Y., and Mr. Christopher Slade, White Hall, Baltimore Co., Md.) and retained there for three months without showing the slightest indication of disease. Later I applied the measure on a number of herds with thoroughly satisfactory results. About ten years later a similar resort was had by Arloing and Rossignol with corresponding success.
Two advantages come from this method: 1st, there is no local infection and no marked swelling so that the injection can be made on the side of the neck where the skin is thin, and clean, and the connective tissue abundant, and where there is less risk of extraneous infection than in the too often dirty or filth soaked tail; 2d, as no living germ is introduced there is no possibility of propagating the disease to other neighboring susceptible animals. Two per cent. of loss, of animals inoculated by the Willem’s method is counted on, but with the sterilized virus there is not even a remote probability of loss. Infection cannot occur from the animal injected with the sterilized virus, so that it can be safely applied among cattle that have not been exposed to infection, but which are likely to be in the future, and these injected cattle can be left to mingle with others that have not been injected without a shadow of danger to either.