Injection of blood vessels may be performed on small animals, or on individual human organs after removal from the body. The object is to fill the vessels with a coloured fluid which will solidify afterwards. It is possible in the same organ to inject the arteries with a red medium, the veins blue, and secretory ducts, such as bile ducts, yellow or blue.

The most convenient basis for an injection mass is gelatine, as its solutions liquefy at a temperature of about 100°F., and solidify a little below that point, and when solidified cut readily, and do not tend to become brittle. The various masses are prepared as follows:—

Red injection mass (Woodhead’s formula) consists of gelatine softened by mixture with water and coloured by carmine.

Dissolve the carmine in the ammonia in a mortar. Pour on the water. Mix thoroughly and filter.

Allow it to stand in the cold water until the water is absorbed and the gelatine has become soft.

Warm (1) almost to boiling point over a Bunsen burner, and add the gelatine slowly. Stir thoroughly and add a ten per cent. solution of acetic acid until the solution becomes slightly acid. This will be shewn by the mass assuming a darker and duller colour. A little salicylic acid may be added to preserve it.

Blue injection mass.—To the gelatine mass (2) prepared as above, and liquefied by heat, add instead of carmine

Every trace of alkali must be kept away from the mass during and after the preparation. Sections of injected organs should be mounted in Farrant’s solution slightly acidulated with formic or acetic acid. With every care, however, the blue colour is apt to fade in the course of time.

Green injection mass. Robin’s formula (modified).

Mix and add one part to three parts of the gelatine mass made as for the red and blue injections.

Method of injection.—In injecting the vessels of tissues it is necessary that the organ or the entire animal, as the case may be, shall be kept during injection at a temperature well above that at which the gelatine mass will melt, otherwise the gelatine will “set” in the arteries and will never reach the capillaries. This warming is effected by immersing the animal in a water bath. The liquefied gelatine is forced into the artery by a syringe or by air pressure. It is essential that the pressure be uniform and steady. This is so much more easily managed with air pressure that this method is strongly recommended to the beginner. But, whatever method be adopted, perfect results can only be obtained with certainty after long practice. Sometimes too high pressure will be employed and the vessels give way, at others the injection may not reach the capillaries at all. The most scrupulous attention to details is essential.

By far the most effective apparatus for injecting is the modification of Ludwig’s constant pressure apparatus devised by Fearnley.2 Although the apparatus appears complicated, the various parts are easily obtained and it would be easy to improvise a substitute for the water bath.

The apparatus which is shewn in figures 10 and 11 consists of a bath deep enough to contain the animal, and a vessel containing the injection fluid. The bath is kept at a temperature of about 110° by an ordinary Bunsen burner. A large Wolff’s bottle (20–40oz.) with three necks, is fitted with three india-rubber stoppers perforated by glass tubes. Through the central stopper a glass tube connected by a rubber tube, with an ordinary Higginson’s syringe, passes almost to the bottom of the bottle. From one of the other necks a rubber tube passes to an ordinary mercurial manometer, while from the third a tube passes to the flask containing the liquefied injection mass, which is immersed in the water bath. This flask is also firmly stoppered, and should be about half filled with injection material. The delivery tube from the large Wolff’s bottle should only just come through the cork. Another glass tube passes down almost to the bottom of the flask, and is connected by a rubber tube with the cannula inserted into the artery. It will be evident from figure11 that when water is pumped by the Higginson’s syringe into the Wolff’s bottle the pressure there will be raised (as indicated by the manometer). This increase of pressure will equally affect the air inside the bottle containing the injection fluid, and the fluid will be forced out along the tube and through the cannula into the artery.

Before using the apparatus a clamp should be placed on the exit tube of the vessel containing the injection fluid, and the pressure should be raised to see that the apparatus is everywhere air-tight. Any leaks should be sealed before the actual injection is commenced.

If an isolated organ is to be injected, a cannula of glass or brass should first be inserted into the artery and securely tied in position. The organs, if cold, must be soaked in water at 120°F. for about half an hour and then transferred to a water bath.

In the case of injecting an entire animal, such as a rabbit, rat, or guinea pig, the injection is best made a few minutes after death. The animal may be chloroformed, and then bled to death by opening a large vein. As soon as death has occurred incise the skin over the thorax in the middle line. Cut through the costal cartilages to the right of the sternum, and through the junction of the manubrium and body of the sternum. These incisions being for most part through non-vascular parts will not lead to escape of fluid during injection. The sternum being forcibly raised towards the left, the pericardium will be exposed and must be carefully divided. An incision must be made into the left ventricle, and a cannula passed up into the aorta and firmly secured by a ligature passed round the aorta with the assistance of forceps or an aneurism needle. Any blood is cleaned away and the animal is then placed in the water bath for about ten minutes. The tube from the bottle containing the injection fluid is then filled by gentle pressure on the syringe, and clamped when full. Its end is then placed on the cannula and secured there by a ligature. The pressure should be raised by squeezing the syringe until the manometer registers one inch. The clamp should then be removed and the injection commenced. The pressure should be raised very gently and constantly by working the syringe, and the condition of the gums, lips, and eyes of the animal observed. The gums will soon shew a pink tinge. The best indications are obtained by watching the effect on the small vessels of the sclerotic. When these are completely filled, which will be in about five to ten minutes according to the rate at which the air pressure has been increased, the injection may be stopped. This result will be obtained, under good conditions, before the manometer indicates a pressure of five inches. The aorta should now be ligatured, and the animal placed in cold water frequently renewed until it is thoroughly cooled. The organs may then be removed and placed in methylated spirit and hardened. Sections are afterwards cut and mounted in the usual way.