216 A MANUAL OF BACTERIOLOGY 



must be pipetted off into a small tube and centrifuged for 

 a few minutes. The suspension must not be too thick, 

 otherwise the leucocytes will take up an unaccountable 

 number of cocci ; the proper density can be judged by 

 experience alone, but the suspension should be only faintly 

 opalescent. Suspensions of pneumococci and other organ- 

 isms are made in the same way. Variations in the number 

 of bacteria ingested may occur according as recently isolated 

 or old strains are employed. 



Instead of centrifuging, the suspensions may be filtered 

 through a double thickness of filter-paper. 



5. Suspension of living leucocytes. To prepare this, 

 take about 10 c.c. of physiological salt solution containing 

 J per cent, of sodium citrate, to prevent the coagulation 

 of the blood. This must be freshly prepared (or kept 

 sterile, which is inconvenient), and the simplest method 

 is to use " soloids " prepared for the purpose by Burroughs 

 and Wellcome ; one of these dissolved in 10 c.c. of distilled 

 water will yield the solution required. This is put into 

 a centrifuge tube and warmed to blood-heat. A healthy 

 person is then pricked in the ear or finger, and his blood 

 is allowed to drop into the fluid until 1 c.c. or more has 

 been collected. The tube is then centrifuged until all the 

 corpuscles have come to the bottom and the supernatant 

 fluid is left clear. If the deposit is closely examined the 

 red corpuscles will be seen to be at the bottom, whilst 

 above them there is a thin grey layer of leucocytes. The 

 whole of the clear fluid is then pipetted off, as close as 

 possible to the leucocyte layer, but without disturbing 

 the latter, with a pipette armed with an india-rubber teat, 

 or with a syringe. The tube is again filled with saline 

 solution, the blood and fluid are mixed, the mixture is 

 centrifuged, and the clear fluid pipetted off, and this 

 process of washing is repeated. Next, the leucocyte layer 

 with the upper layer of red corpuscles (which also contains 



