o88 ' H. B. Seddon .- 



Note: — Set IV., ut the end of 24 hours' incubation, showed 

 positive agglutination with 0.05 c.c. of emulsion — i.e., in Tube A, 

 but not in the other tubcvS — B, (', and D. 



In tubes B and C, containing O.OL'5 and 0.01 c.c. emulsion, a 

 positive agglutination was manifest at the end of three days' in- 

 cubation, but the deposit in tube D, with 0.005 c.c. emulsion, even 

 at the end of this time, was very small indeed — in fact, barely 

 appreciable. 



The controls, it should be mentioned, i-emained unchanged. 



From these experiments it appears as if the large volume of 

 Huid (20 c.c.) in Set IV. affected the rate of sedimentation of the 

 clumps of bacilli in those tubes with the smaller quantities of 

 emulsion. As, however, such a large quantity of fluid is not used 

 in the tubes in making a diagnostic test, these results in Set. IV. 

 are not of great impoitance. In the practical application of the 

 test the total volume of fluid in each tube is usually made up to 

 about 1.5 c.c. A number of tests have also been made in which 

 the total amount was 2 c.c. The tubes in Sets 1, 2 and 3, where 

 the total volume has been made up to 2 c.c, are, therefore, of 

 primary importance. Here it is found that such a quantity of 

 emulsion and of total fluid have been employed that the complete 

 agglutination reaction takes place within the first 24 hours. 



Tubes of 2.5 c.c. capacity have been found very suitable in mak- 

 ing the test, and the amounts of diagnostic material (serum, whey, 

 etc.), and of emulsion, are, even over a lai-ge range of quantities, 

 but involving only a few different basal dilutions, easily contained 

 in 1.5 or 2 c.c. of fluid. Also, 24 hours is a convenieTit time for 

 incubation l)efore reading the results. 



Where the total volume of Huid in each tuljc is niade up to 1.5 

 or 2 c.c, with incubation extending over 18 to 24 hours, the 

 optimum of emuhioti will he the smallest quontiti/ which gives re- 

 sults thitt are easily read, wiietlier the agglutination teactioii be 

 positive or negative. 



