New York Agricultural Experiment Station. 83 



a platinum loop of such a size that it will transfer an average of 

 0.004 gram of the stained milk. The area over which the mixture 

 spreads is determined by a cover glass 18 millimeters square. The 

 counting of the bacteria is done immediately without allowing the 

 film to dry. 



Donald 13 has made use of the microscope in the bacterial analysis 

 of distilled water. He has devised an ingenious scheme for making 

 pipettes which will yield uniform and accurate drops of any desired 

 size. This method of measurement, where it can be used, will 

 improve the accuracy of measurement employed in the direct 

 microscopic method of counting bacteria, but is a more complicated 

 procedure than measurement by means of capillary pipettes. 



TECHNIQUE. 



Microscopical method. — The microscopical method used in this 

 work consisted in measuring out 0.01 cubic centimeter of milk taken 

 directly from a well-shaken sample by means of a specially graduated 

 and accurately calibrated pipette (See page 105). The drop of milk 

 was deposited on a clean glass slide and spread over an area of one 

 square centimeter with a .stiff, straight needle. Duplicate smears 

 of each sample were made on the same slide. The milk was then 

 dried by gentle heat which was obtained by means of a level wooden 

 surface over a steam radiator. Care was exercised not to allow the 

 smears to become too hot as this made them check and thereby 

 made satisfactory staining impossible. As soon as dry, the slides 

 were placed for a short time in a Coplin staining jar containing 

 xylol to remove the fat. They were then taken out and the surplus 

 xylol about the edges of the slide wiped off with filter paper. The 

 smears were dried and then fixed to the slide by means of a 95-per-ct. 

 solution of alcohol. Immediately thereafter they were stained 

 from two to three minutes in Loeffler's methylene blue, after which 

 they were decolorized to a light blue in a 95-per-ct. solution of 

 alcohol. 



The counting was done under a 1.9 millimeter oil-immersion lens. 

 With the particular microscope used the draw tube was placed at 



18 Donald, R. An apparatus for liquid measurement by drops and applications in 

 counting bacteria and other cells and in serology, etc. Proc. Roy. Soc, Ser. B, 

 86 : 198-202. 1913 : A method of counting bacteria in water. Lancet, 184 : 1447- 

 1449. 1913. 



