410 MILK AND MILK PRODUCTS 
The number of ccllular clements per cubic millimeter of milk= 
56,000y 
lid? ’ 
of squares which just spans the diameter. d is determined once for all 
by marking the ricroscope draw tube so that only 20 fields have to be 
counted, and the figures substituted in the formula. 
Doane-Buckley Quantitative Method for Estimating Leucocytes in 
Milk. With this method 10 ¢.c. of milk are centrifuged for four min- 
utes in graduated sedimentation tubes, at an approximate speed of 
2000 r.p.m. The cream is lifted out with a cotton swab, care being taken 
to get as much as possible of the fat. It is then centrifuged one minute 
more and the cream again removed with a cotton swab. Any fat re- 
maining in the milk interferes seriously with the counting, as, if there 
are more than a few globules they form a layer on the top of the liqiuid 
in the counting chamber, and, as the leucocytes settle to the bottom of 
the chamber, it is difficult to see through the fat. It is only with cows 
giving milk difficult of separation where this trouble is experienced, and 
with such animals considerable care is necessary in removing all the 
cream gathered at the top of the sedimentation tube. The method of 
removing the fat with cotton is the best one that has occurred to us, and 
it is the only part of the process that does not operate with entire satis- 
faction in every instance. 
Following the removal of the cream, after the second centrifuging, the 
bottom of the tube will contain a portion of the sediment which is easily 
seen. This sediment may, in extreme cases of cows suffering from 
garget amount to as much as 1e.c. Ordinarily it will be considerably 
less than $ c.c. The amount varies considerably with the number of 
leucocytes, but not absolutely. The milk above this sediment is 
removed with a small siphon, whith can be easily arranged with bent 
glass tubes drawn to a fine point and supplied with a small rubber end 
pinch cock. In using the siphon it 1s better to keep the point near the 
surface of the milk in the tube in order not to agitate the precipitated 
leucocytes and draw a number of them off with the milk. The milk in 
the tube may be siphoned within an eighth of an inch of the sediment 
in the tube. This will usually be below the 3-c.c. mark. Two drops of 
saturated alcoholic solution of methylene blue are then added, thor- 
oughly mixed with the sediment by shaking, and then set in boiling water 
two or three minutes to assist the leucocytes in taking the color. The 
contents of the tube can be boiled by holding it directly in the flame, but 
it has no advantage over the use of the water bath, and it is very likely 
to break the glass. After heating, some water is added to the tube to 
where y= the average number per field of vision, d=the number 
