44 VACCINE AND SERUM THERAPY. 



mentioned before, there are 'conflicting views concerning the effect 

 of dilution of serum for which the opsonic index is to be 

 determined. 



DIFFICULTIES ARISING BECAUSE OF THE PARTICULAR 

 SPECIES OF BACTERIA IN THE SUSPENSION. 



Anyone who has made opsonic index determinations has at 

 times had slides to examine in which the bacteria were not 

 well stained and in which the microorganisms were apparently 

 clumped. Frequently only shells of organisms remain and at 

 times the bacteria are completely dissolved. These difficulties 

 are apparently dependent upon the particular culture used but 

 to a greater extent upon the agglutinins and lysins present 

 in different sera. To overcome this difficulty young cultures 

 are preferred because they stain better than do old cultures. 

 To overcome agglutination and lysis the serum is in some cases 

 heated to 55 or 65. The clumping of bacteria in opsonic 

 index preparations is given little attention by Wright. Other 

 investigators have, however, regarded clumping of bacteria 

 as responsible for the occasional enormous differences of the 

 indices determined. Walker gives tables which indicate the 

 objection of clumps on the uniformity of phagocytosis and has 

 prepared a technique by which he obtains clump-free bacterial 

 suspensions. According to this technique the bacteria are rub- 

 bed gently in a mortar with small, and gradually increasing, 

 quantities of salt solution until a thick suspension is obtained. 

 This suspension is then filtered through moistened scraped filter 

 paper until in a stained specimen no more -clumps are found. 

 Even though a technique may be devised to obtain relatively 

 clump-free suspensions of bacteria, this will not do away with the 

 clumping occasionally observed in opsonic index preparations, 

 because the action of agglutinins in undiluted or only slightly 

 diluted sera will again cause clumping of the bacteria. 



LEUCOCYTES. 



There -has been but little criticism of the method of collection 

 of leucocytes, as proposed by Wright. Simon has used 0.1 per 

 cent ammonium oxalate instead of 1 . per cent sodium citrate 

 in salt solution. Some investigators have drawn the corpuscles 



