22 J. PERCY BAUMBERGER 



The growth of larvae on yeast nucleoprotein media is shown in 

 figure 7 and table 7. Table 7 shows that larvae do not grow on 

 nucleoprotein in the absence of Pasteur's solution, but, if these 

 salts and sugars are added, do grow rapidly and fomi large numbers 

 of nonnal pupae from which normal highly fertile adults emerge. 



yeast) were killed by ether and broken open by grinding in a mortar with a quan- 

 tity of pure white diatomaceous earth, adding enough water to keep the mass in 

 a sticky, smooth condition. The grinding was continued till examination with a 

 1.6-mm. objective showed that many of the yeast cells had been cut into irregular 

 rectangles. The yeast was then poured with the addition of 0.4 per cent MaOH 

 solution into a large bottle and 8000 cc. of the alkali were added. About 40 cc. of 

 chloroform were then mixed with the solution to prevent the development of bac- 

 teria. The contents of the bottle were thoroughly agitated several times a day. 

 After forty-eight hours the supernatant fluid was poured off, leaving a great part 

 of the yeast and diatomaceous earth in the bottle. This fluid was centrifuged in 

 a large-sized electric centrifuge with a capacity of four 250 cc. bottles. Each liter 

 was run for twenty minutes (fifteen minutes at maximum speed) and the super- 

 natant fluid carefully poured off. This fluid was examined for yeast cells with a 

 1.6-mm objective and showed an entire absence of them. After all the yeast cells 

 had been removed in this manner, the liquid had a clear opalescent color and 

 proved to be rich in nucleoprotein. This was precipitated in great white floccules 

 by adding 10 per cent HCl in drops. The precipitate dissolved in the alkaline or 

 neutral solution, but remained in the slightly acid solution in which the largest 

 amount was formed. The precipitate was separated pure from the solution by 

 centrifuging and washing with acid alcohol and neutral alcohol in which it was 

 insoluble. The white precipitate was then dried over H2SO4, forming a white 

 powder. The remaining fluid was neutralized with N/10 NaOH and dialyzed for 

 five days in running water, keeping the surface covered with toluol. No precipi- 

 tate was formed. The neutral, salt-free solution was then heated to boiling and 

 acidified with a drop of HCl or acetic aid and also acidified and then boiled. No 

 marked precipitate was formed. Half saturation, complete saturation with 

 (NH2)S04 when hot or cold and saturation with NaCl and with picric acid failed 

 to bring down any precipitate. A heavy precipitate which appeared to be a pep- 

 tone decomposition product of the nucleoprotein was produced upon the addition 

 of phosphomolybdic acid. 



The nucleoprotein was ground in a mortar and then made into media as fol- 

 lows: 1) Nucleoprotein moistened with tap-water was placed in test-tubes and 

 sterilized in the autoclave; 2) nucleoprotein moistened with Pasteur's nutrient 

 solution (grape-sugar, cane-sugar, ammonium tartrate, MgS04, K2HPO4, HoO) 

 in test-tubes and sterilized; 3) nucleoprotein and 1.5 per cent agar-agar tap- 

 water solution autoclaved and mixed aseptically in sterile test-tubes, and, 4) 

 nucleoprotein. Pasteur's nutrient solution and 1.5 per cent agar solution 

 autoclaved and mixed aseptically in sterile test-tubes. If mixture is made before 

 autoclaving the protein adsorbs (?) the agar and on cooling jellation does not 

 take place. 



