METHODS OF STUDYING H.EMOLYSINS. 329 



for activation. Most of the complements will keep unchanged for 

 a number of days provided the serum is kept on ice. But this does 

 not preclude unpleasant surprises, diminutions in the complementing 

 power often occurring to a high degree without any assignable cause. 

 According to our experience the complements of guinea-pig serum 

 and goat serum are relatively stable. The least reliable in this respect 

 is horse serum, whose complementing powers are often partially or 

 completely destroyed within twenty-four hours. The complements 

 also suffer when the serum is dried : at least that has been the case in our 

 rather limited experience. 



The best method of preserving the complements for a long time, 

 and the one almost always reliable in all cases, consists in freezing 

 the serum at --10 to -15C. This method has been employed in 

 the Institute for a long time. The serum is bottled in little vials, 

 which are then kept in a freezing apparatus or in a well-insulated 

 freezing mixture of ice and salt, each vial being thawed out as needed. 

 This procedure is at present the only one which is of general appli- 

 cability and w r hich preserves the various constituents of the serum for 

 a long time. 



The blood used for the hffimolytic tests is defibrinated by one of 

 the methods above mentioned. In special cases, instead of defibrina- 

 ting, one can prevent coagulation by precipitating the lime salts. 

 This is done by allowing the blood to flow into salt solution to which 

 citrate of soda has been added, as was recommended by Ehrlich. 1 

 For the majority of experiments the blood is diluted with physiological 

 salt solution. If for any reason one wishes to remove the serum, the 

 blood is separated by centrifuge and the suspending fluid renewed 

 several times. As a rule blood which has been kept on ice for two 

 days can still be used. 



It should also be mentioned that a suitable salt solution should 

 be employed for each species of blood. For the blood-cells of most 

 mammals a feebly hypertonic solution of NaCl 0.85% is best adapted. 

 In 0.85% salt solution dog and horse blood frequently shows a slight 

 amount of spontaneous haBmolysis which can often be prevented by 

 using a somewhat higher concentration (0.95%) of the salt. As 

 a rule strongly hypertonic solutions of salt are to be avoided because 

 the increased contents of salt markedly inhibits hamolysis. 2 



1 Ehrlich, Fortschritte der Medizin, 1897, No. 2. 



2 S. Markl, Zeitsch. f. Hygiene, Vol. 39. 



