HEMOLYSIN8. 257 



undiluted blood is centrifugated, the overlying 

 fluid drawn off by means of a pipette and substi- 

 tuted by fresh salt solution; the corpuscles are 

 thoroughly mixed with the new solution and the 

 process of centrifugation repeated, the corpuscles 

 finally being diluted to the original volume with 

 salt solution. After from two to four washings 

 any residual serum usually may be disregarded. 

 To test the hemolytic power of a serum one meas- 

 ures identical quantities of the 5 per cent, washed 

 blood into each of a series of test-tubes by means 

 of a graduated pipette and then adds increasing 

 quantities of the serum to succeeding tubes. All 

 tubes are then diluted to equal volumes by means 

 of salt solution, as it is of some importance to 

 maintain a uniform concentration of the cor- 

 puscles. The contents of the tubes are mixed 

 evenly by shaking and the series is placed in the 

 thermostat for about two hours; this temperature 

 is necessary for complete and rapid action of the 

 toxic substances. At the end of this time the tubes 

 are placed in the ice chest and left over night in 

 order that the cells may settle to the bottom, or 

 sedimentation may be accomplished at once by 

 centrifugation. 



In either case, the overlying fluid is colored red 

 by the dissolved hemoglobin in proportion to the 

 extent of destruction of the erythrocytes. In case 

 solution has been complete, the sediment is indis- 

 tinct and colorless, being made up only of the 

 stromata of cells, whereas in the tubes showing 

 only partial hemolysis the sediment is red and has 

 an indirect quantitative ratio to the coloration of 

 the overlying fluid. By suitable variations in the 

 amounts of serum used in different tubes, its 



