1 88 PRACTICAL METHODS IN IMMUNITY 



particular organism and not for the other organism. This variation may be of 

 the nature of a high or low opsonic index. 



METHOD OF WRIGHT FOR OBTAINING OPSONIC INDEX 



While other observers had previously noted the presence of sub- 

 stances in immune sera which so acted on the bacteria that phagocytosis 

 was made possible, yet it was to Wright and Douglas, in 1903, that the 

 existence of this factor in phagocytosis was brought forward and the 

 estimation of such substances made practicable. 



To this substance the name opsonin was given the Greek word from which it is 

 derived indicating preparation of the food that is, the opsonin so alters or sensitizes 

 the bacteria that they can be engulfed or phagocytized by the polymorphonuclear 

 leukocytes (the microphages of Metchnikoff) . About the same time Neufeld and 

 Rimpau noted the presence of a substance in immune sera which so acted on bacteria 

 as to prepare them for phagocytosis. Their designation "bacteriotropic substance" 

 is practically synonymous with opsonin. 



In 1902 Leishman introduced the method of determining the "phagocytic index." 

 By taking i part of blood and impart of an emulsion of the bacteria in question and 

 keeping the mixture in a moist chamber at body temperature for a standard time, 

 as fifteen to thirty minutes, and then spreading the blood-bacteria mixture and 

 staining the film with Leishman or Wright's stain he counted the number of bacteria 

 in a certain number of polymorphonuclears, and by dividing obtained the average 

 number per leukocyte of bacteria phagocytized. 



The Wright technic for determining the phagocytic average, and 

 from this the opsonic index, is as follows: 



Blood is taken from the patient and at the same time from a normal individual, 

 or preferably the blood of several normal individuals is pooled. This blood is best 

 collected in a Wright's tube, although it may be received in a small test-tube. After 

 coagulation and separation of the serum, the serum is ready for use. 



The next step is to prepare the leukocyte gmulsion. For this we fiU a centri- 

 fuge tube with normal saltsolution, to which has been added i% 'sodium, citrate 

 the latter to prevenTcoagulation. Then having pricked a finger congested by a 

 constricting rubber band, from 15 to 20 drops of blood are added to the citrated 

 salt solution, and the mixture thoroughly shaken. After centrifugalization for about 

 five minutes the red corpuscles will be thrown to the bottom of the tube with the 

 leukocytes forming a superimposed layer. In order to free the leukocytes entirely 

 from serum admixture, the supernatant citrated salt solution is pipetted off, and a 

 fresh tubeful of salt solution is added to the blood-cell sediment. Again shak- 

 ing, we centrifuge, obtaining for a second time a sediment of blood cells with the 

 leukocytes in the superimposed layer. In some laboratories the washing in salt 

 solution is again repeated, but for all practical purposes two washings as described 

 above suffice. 



The superimposed layer of vyhite cells may now be pipetted off from the heavier 



