TUBERCULOSIS. 219 Kitasato's method of washing the sputum has been modified and simplified by Czaplewski and Hensel.1 In their studies of whooping-cough, instead of washing the flakes in water in dishes, they shook them in peptone water in test-tubes. The shaking in the test-tube being so much more thorough than the washing in dishes, fewer •'changes are necessary, three or four washings being sufficient. In 1887, Nocard and Roux gave a great impetus to investigations upon tuberculosis by their discovery that the addition of 4-8 per cent, of glycerin to bouillon and agar-agar made them suitable for the development of the bacillus, and that a much more luxuriant development could be obtained upon these media than upon blood-se- rum. The growth upon such '' glycerin agar-agar'' (Fig. 62) very much resembles that upon blood-serum. The growth upon bouillon with 4 per cent, of glycerin is also luxuriant. As tubercle bacilli require considerable oxygen for their proper devel- opment, they grow only upon the surface of the bouillon, where a rather thick myco- derma forms. The surface- growth is rather brittle, and after a time gradually sub- sides fragment by fragment. The tubercle bacillus can be grown in gelatin to which glycerin has been added, but as its development takes place only at 37°-38° C., a temperature at which gelatin is always liquid, its use for the purpose is disadvantageous. 1 Centralbl.f. Bakt. z/. Parasitenk., xxii., Nos. 22 and 23, p. 643. FIG. 62.—Bacillus tuberculosis on "glycerin agar-agar."