232 



SCIENCE. 



[Vol. XXII. No. 560 



elaborate antitoxins, and if so, what can be said about the 

 natnre of these antitoxins'? These are q:iestions in which 

 I have been deeply interested for some time, and which I 

 have attempted to solve. In this attempt, I have born in 

 mind the fact that these organs are the source of the nuc- 

 leated white blood coriDuscles. Do these corpuscles con- 

 tain a germicidal or antitoxie substance, and if so, what is 

 its nature? The chief chemical constituent of nuclei is a 

 substance called nuclein, some of the general properties of 

 which are known to ^physiological chemists. Can it be 

 that nuclein is the germicidal or antitoxic substance? 

 Have the nucleins in general or as a class any germicidal 

 action? As methods of insolating the nuclein are known, 

 these questions can be answered by experimentation, and 

 this I have attempted to do. 



At first I tried to prepare an active nuclein from com- 

 pressed yeast, but the results were not satisfactory. Com- 

 pressed yeast contains a large amount of water and starch. 

 The large pro^Jortion of the first mentioned constituent 

 caused a very small yield of nuclein, and there were many 

 dffficulties in the complete separation of the starch. 

 There were, however two other and more serious objec- 

 tions to the use of compressed yeast. The first of these is 

 due to the fact that such yeast contains bacteria to begin 

 with, and the nuclein contained in this yeast has already 

 been decomposed. The second difficulty lies in the fact 

 that comj)ressed yeast contains many dead cells, and an 

 active nuclein can be obtained only from living, healthy 

 cells. 



From the cells obtained from pure cultures of yeast, I 

 have obtained an active nuclein by the following method: 



The cells from j)ure cultures of yeast are washed with 

 sterilized water, then treated with a five per cent solution 

 of potassium hydrate and filtered thi-ough paper. Steriliz- 

 ation of the jDaper is not necessary. The filtrate is feebly 

 acidified with hydrochloric acid and the proteid precipi- 

 tated with 96 per cent alcohol. The precipitate is washed 

 with alcohol by decantation until the supernatant fluid 

 remains colorless. The precipitate is then collected upon 

 a filter, and after all the alcohol has passed through, it is 

 dissolved in very dilute potassium hydrate (.25 to .50 per 

 cent). This inquire nuelein h^s marked germicidal effects 

 upon the stajjhylococcus pyogenes aureus, albus, the an- 

 thrax bacillus, and the germs of typhoid fever, Asiatic 

 cholera, and tuberculosis. 



The following experiment will illustrate the action of 

 this nnclein upon the bccillus of tuberculosis: A loop of 

 tuberculous si^utum, showing from 40 to 60 bacilli in each 

 field when stained, was stirred up in beef tea, allowed to 

 stand for twenty-four hours at 38" C. and injected into the 

 abdominal cavity of guinea pig No. 1. Another loojJof the 

 same sputum was added to a solution of 30 milligrams of 

 inpure yeast nuclein in .08 per cent of jjotassium hydrate, 

 and this was allowed to stand in the incubator at 38 de- 

 grees C for twenty-four hours, and then injected into the 

 abdominal cavity of guinea i^ig No. 2. 



At the expiration of fourteen days, both of those ani- 

 mals were killed. The omentum of No. 1 was a tubercu- 

 lous mass throughout, while No. 2 showed not the slight- 

 est evidence of the disease. 



I have isrepared testicular nuclein from the testicles of 

 of the bull, dog. guinea pig, and rat. The testicles are 

 stripped of their investing membranes as soon as removed, 

 rubbed u.p and extracted rejjeatedly with a mixture of 

 equal volumes of absolute alcohol and ether. Then, the 

 testicular substance is digested for some days (until the 

 supernatant fluid fails to resjjond to the biuret test for 

 peptones) at 40 degrees C. with pepsin and .2 per centhy- 

 drocoloric acid. The undijested portion which contains 

 the nuclei is collected on a filter paper and washed, first 

 with .2 per cent hydrochloric acid, then with alcohol. Fi- 



nally it is dissolved in a .5 per cent solution of potassium 

 hydrate and filtered through a Chamberland filter without 

 pressure This solution is clear, more or less yellow, and 

 feebly alkaline. On the addition of nitric acid, a 

 white precipitate forms and dissolves colorless in the 

 cold on the further addition of nitric acid. This nu- 

 clein does not give the biuret reaction, but does respond 

 to the Millon test. The nitric acid solution of the preci- 

 pitate becomes yellow on the addition of ammonia. This 

 nuclein also has germicidal properties, as is demonstrated 

 by the following exj)eriment: 



A solution of testicular nuclein of unknown strength, 

 obtained from the testicles of a bull, was diluted with 

 four volumes of physiologic salt solution, inoculated with 

 the bacillus anthracis, and plates made with the following 

 I'esults; 



Time, Immediate. 30 min. 1 hr. 2 hrs. 3 hrs. 



Number of colonies, 730 6 



Other nucleins with germicidal properties have been 

 obtained from the thyroid gland, spleen, and from the 

 yolks of eggs. 



These experiments render it highly probable that the 

 nuclein-forming organs of the body have some concern in 

 the production of immunity. The nucleins formed by 

 these cells or in these organs pass into the blood partly 

 in the form of multinuclear white corpuscles — the so- 

 called phagocytes. 



In order to state my views upon immunity in a con- 

 densed form, I will summarize as follows: There must 

 be three factors in the production of immunity in an ani- 

 mal naturally suscej)tible: First, there must be an in- 

 citing or immunizing substance introduced into the body. 

 This substance is the nuelein of the germ. These nu- 

 cleins, when introduced into the bodies of certain ani- 

 mals, in certain amounts and under certain conditions, 

 have the jjroperty of so stimulating the activity of certain 

 organs that those organs produce and supply to the 

 blood an antidote to the substance introduced. 



Secondly. The organs whose activity is stimulated by 

 these immunizing agents are those such as the spleen, 

 thyroid gland and bone marrow, which manufacture nu- 

 cleins. 



Thirdly. The antidotal substance is a nuclein. The 

 kind and amount of nuclein formed will depend upon 

 the nature of the inciting agent and the condition of the 

 organ or organs acted upon. 



I use the word "nuclein" in a broad sense, including the 

 true nucleins, nucleinic acid, and^nucleo-albumins. By the 

 term "nuclein" I mean that part of the cell which under 

 normal conditions is endowed with the capability of growth 

 and reproduction, which assimilates other proteids and 

 endows these assimilated substances with its own j)roper- 

 ties. It is that part of the cell which gives in its indivi- 

 duality. Whether these nucleins while in solution and 

 devoid of morphologic unity are still capable of assimilat- 

 ing allied bodies cannot at present be satisfactorily deter- 

 mined. 



We can suppose that the process of immunizing an ani- 

 mal proceeds in something like the following manner : 



The modified virus of tetanus, is introduced into some 

 distant j)art. In some unknown way, the spleen is stimu- 

 lated to action and secretes a nuclein which is carried par- 

 tip in solution, partly in the form of multinuclear cells, to 

 the invaded party of the body, and the tetanus poison is 

 converted into the nuclein coming in contact with it, or is 

 otherwise rendered inert. Later, a larger quantity of the 

 tetanus poison is introduced, and now the acspleeu ts more 

 promptly and energeticallj' than before. This prompt- 

 ness and energy of action are increased by exercise, and 

 finally an amount of tetanus culture of full virulence, suf- 



