﻿340 The Philippine Jout-nal of Science isu 



FIXATION AND STAINING 



Fixation and staining can be carried out in hanging drops 

 with the pipette. The stain or fixing reagent may be placed 

 under the cover with a platinum loop or coarse pipette and 

 applied with the isolating pipette to the organism. The medium 

 in which the cell lies may be withdrawn and the cell washed, 

 following the method outlined for freeing cells from bacteria 

 or serum (see page 337). The fixative, and then the stain, 

 may be applied with the pipette. The same pipette may be 

 used for both by washing it between each use with water or 

 other fluid in hanging drops. If the fixative is volatile or can- 

 not be easily washed out of the pipette, one may keep it in a 

 separate pipette. A second holder may be used or the pipette 

 removed from the holder and the one containing the stain 

 substituted. After staining, the cell may be washed and de- 

 colorized if necessary. If it is desired to mount the stained 

 preparation in 'balsam, albumen or other fixative may be ap- 

 plied and the location of the cell marked by Brunswick black 

 or India ink on the upper side of the cover. The preparation 

 is then dehydrated by drying on alcohol and mounted. Or the 

 cell or colony may be located, and the cover removed, dried, and 

 stained. In Plate I, figs. 3-7, a photomicrograph is shown of 

 a preparation of Bacterium coli. By means of successive iso- 

 lations, a series of 5 droplets was obtained, containing respec- 

 tively 1, 2, 4, 8, and 16 bacilli. The preparation was dried in 

 toto, stained, washed, and mounted without the loss of a single 

 bacillus in the series. I have also used this method in staining 

 young colonies of Bacillus tuberculosis derived from an isolated 

 single cell. Drops of different stains may be dried on a large 

 cover. When they are to be used, the cover is placed over 

 an isolating chamber, and the stain, dissolved in the water of 

 condensation, is taken up with the pipette. By changing covers 

 and using two pipettes, one may obtain a great variety of 

 combinations. 



SEROLOGICAL TESTS 



I have tested only agglutination and precipitation by the pipette 

 method. The agglutination in hanging drop is especially practi- 

 cable with motile bacteria rapidly agglutinated, such as vibrios of 

 Asiatic cholera and typhoid bacilli. The serum in proper dilution 

 is taken into a pipette and applied to the edge of a hanging drop 

 containing the bacteria. This method gives very striking results 

 with vibrios. When a very small quantity of cholera agglutinat- 

 ing serum is applied at the edge of an emulsion of cholera vibrios, 



