6 THE AMERICAN MONTPILY [January, 



spread on a level tray. On each cover-glass is placed, by means of a 

 flamed pipette, a moderately large drop of sterile water (distilled or 

 h)drant). This will spread over the entire surface of the cover if it 

 has been properly cleaned. The end of a flamed platinum wire is very 

 gently touched to the surface growth of an agar or gelatine culture of 

 tlie germ in question, after which it is very carefully immersed two or 

 three times in as manv places in the water on each cover-glass. A 

 sufticient number of bacteria will adliere to the end of the wire to 

 make from four to ten preparations. The cover-glasses are then placed 

 in an incubator at a temperature of about 36° C, where they are al- 

 lowed to remain until the water is evaporated. 



Many of the bacteria by means of their power of locomotion will 

 become separated from the clump of germs introduced by the wire, and 

 will be found on the drying of the preparation to be distributed very 

 satisfactorily arovmd these centers. Tliis natural separation of the bac- 

 teria prevents the breaking oft' of the flagella by stirring or other arti- 

 ficial means employed in securing the necessary distribution and isola- 

 tion. In this manner I have isolated the bacteria in a preparation so 

 that in many fields not more than a score of germs could be seen, and 

 the excellent condition of their flagella warrant my recommending this 

 method of preparation. The fact that the bacteria are properly isolated 

 over only a small portion of the preparation is not necessarily an objec- 

 tion. Our hopes, however, are not yet fully realized, for there will gen- 

 erally be a few, often many, detached flagella lying between the bacteria, 

 some of which are partially or wholly deprived of their appendages. I 

 have frequently observed clumps of flagella which gave the appearance 

 of those belonging to an individual germ, the body of which had disap- 

 peared. As these were found in preparations made from old cultures, 

 they suggest the possibility that the body of the germ is first to degen- 

 erate, or else that through some physiological process the flagella are 

 detached»from the bodies of the bacteria after a certain age is attained. 

 These hypotheses are strengthened somewhat by the fact that in older 

 Cultures there appear to be a greater number of detached flagella. 



(2) The Composition of the Mordant. — The mordant recom- 

 mended by Loeffler seems to be the most satisfactory of any thus far 

 suggested for general use. I have found, however, that a mordant 

 which contains only 10 per cent, tannic acid can occasionally be used 

 with advantage. It can be more easily and thoroughly removed from 

 the specimen, and consequently the formation of a troublesome precipi- 

 tate with the staining fluid occurs less frequently. It was found that 

 with the hog cholera bacillus it gave equally as good results as the 

 one containing 20 per cent, tannin ; with the tvphoid bacillus it was 

 not so satisfactory, and with a water bacillus a 20 per cent, tamiin so- 

 lution in the mordant was necessary to secure the staining of the fla- 

 gella. From a limited number of experiments it seems quite probable 

 that a variation in the quantity of the tannic acid in the mordant may 

 be of much service in staining the fiagella on certain bacteria where dif- 

 ficulties are experienced with the use of the mordant prepared after 

 Loefller's formuke. Althougii I have tried a considerable number of 

 the "■ fixing agents,'' I have thus far been unable to stain the flagella 

 with tlie use of any mordant not containing tannic acid. 



In applying the mordant I have met with better results by allowing 



