ESTIMATION OF OPSONIN 39 1 



5. Mark the pipette (with the grease pencil) with the 

 distinctive number of the serum and place it in the 

 glass box or tray. 



6. Take another similarly prepared pipette and aspi- 

 rate into it equal volumes of washed cells, bacterial 

 emulsion and pooled serum. Treat precisely as in 3 

 and 4, label it "control" or "N.S." (normal serum) 

 and place in the box by the side of the specific serum 

 preparation. 



7. Place the box with the pipettes in the incubator 

 and set the signal clock to ring at 1 5 minutes (or start 

 the stop watch). 



6. At the expiration of the incubation time remove 

 the pipettes from the incubator. 



9. Cut off the sealed end of the specific serum prepa- 

 ration. Mix its contents thoroughly as in step 3, and 

 then divide the mixture between two 3 by i slips and 

 carefully spread a blood film (vide page 376) on each 

 in such a way that only one-half of the surface of each 

 slide is covered with blood the free edge of the blood 

 film approximating to the longitudinal axis of the slide. 



Allow films to dry and label the slides with writing 

 diamond. 



10. Treat the contents of the control pipette in 

 similar fashion. 



11. Select the better film from each pair for fixing 

 and staining. 



12. Fixing and staining must be carried out under 

 strictly comparable conditions, and to this end the 

 slides are best handled by placing in a glass staining 

 rack which can be lowered in turn into each of a series 

 of glass troughs containing the various reagents (Fig. 

 196) . Place the rack in the first trough which contains 

 the alcoholic solution of Leishman's stain for two min- 

 utes to fix. 



Transfer to the second trough containing the diluted 

 stain for ten minutes. 



