Differential Counting of Bacteria 



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143 (2103) 



Differential counting of living and dead cells of bacteria. 



By ARTHUR T. HENRICI. 



[From the Department of Bacteriology and Immunology, the Uni- 

 versity of Minnesota, Minneapolis, Minnesota.] 



By means of the procedure here described it is possible to 

 determine with considerable accuracy the number of bacterial 

 cells in a suspension and at the same time to determine the size 

 and form of the cells and to differentiate the living and dead 

 cells. The method makes use of the principle of the Breed and 

 Brew 1 method of counting bacteria in milk, and the negative 

 staining method of Benians. 2 



A measured quantity of bacterial suspension is mixed 

 thoroughly with an equal quantity of 2 per cent, aqueous Congo 

 red solution ; the mixture is allowed to stand ten minutes. After 

 again shaking the mixture, 0.01 c.c. is removed by means of a 

 capillary pipette of that capacity and discharged on to a clean 

 slide which has been clamped to the table over a piece of white 

 paper on which a 2 cm. square has been ruled. By means of a 

 stiff wire the drop of liquid is spread as evenly as possible over 

 this area. After it has become thoroughly dry the slide is im- 

 mersed a moment in a 1 per cent, solution of hydrochloric acid 

 in 95 per cent, alcohol. This turns the dye blue and also fixes 

 the film. If covered with a layer of cedar oil the slides will 

 keep indefinitely, but if exposed to the air they fade considerably. 



Cells which were alive at the time of staining are unstained 

 and appear as white spots on a blue ground. While the cells 

 themselves may shrink considerably after fixation and drying, 

 a number of comparative measurements have shown that the 

 clear space in the film faithfully reproduces the size and form of 

 the living wet cells. With favorable material flagella may be 

 demonstrated by this stain. 



Seiffert 3 has observed that when bacteria are suspended in a 

 weak Congo red solution and examined in hanging drops that 



1 Technical Bulletin No. 49, New York Agricultural Experiment Station. 



2 British Medical Journal, 1917, ii, 722. 



3 Centralbl. fur. Baht., etc., Abt. I, Orig., 1922, lxxxviii, 151. 



