A STUDY OF FIXATION REACTIONS. 359 



be serum obtained from guinea pigs on the day of the experiment; the serum 

 must be free from haemoglobin and must be centrifugated until perfectly clear. 



ft. ANTIGEN. 



Wassermann (2) uses extracts in salt solution from various organs of infants 

 who have died of congenital syphilis, made by taking 1 gram of organ, grinding 

 thoroughly in a mortar with 4 cubic centimeters of 0.85 per cent salt solution, 

 plus 0.5 per cent carbolic acid, shaking for twenty-four hours, and eentrifugating 

 until the fluid is perfectly clear. He uses 1 cubic centimeter of a 0.1 dilution 

 of this extract for each test tube. 



Meier (3) employs only extracts of livers from syphilitic foetuses, as other 

 organs are less active. 



Morgenroth and Stertz(5) suggest that the luetic organs be frozen and kept 

 solid until ready for use, and that from these frozen organs one gram be used 

 for each experiment in preparing a fresh extract. 



Marie and Levaditi(6) thoroughly desiccate the foetal organs and make a 

 powder which retains its strength for months, and for each experiment make up 

 a fresh extract with a little of this powder and salt solution. 



The extract of Wassermann is fit for use for about six days, although Meier(3) 

 finds that an ordinary aqueous extract retains its strength for four months if 

 protected from heat, light and air, and if not too frequently centrifugated. 



C. THE ANTIBODY. 



The antibody is usually obtained from the serum of a patient suspected of 

 suffering with the disease. In a large number of cases the cerebro-spinal fluid 

 has been employed as antibody. Meier (3) draws 6 or 8 cubic centimeters of 

 blood and as quickly as possible obtains at least 1.4 cubic centimeters of serum 

 which is inactivated at once. In the case of children, less blood is drawn, also 

 a smaller quantity of spinal fluid is used. He recommends that the serum or 

 spinal fluid be used immediately because of the danger of rapid deterioration 

 upon standing. In establishing the value of the sero-diagnosis of syphilis Wasser- 

 mann(2) uses the serum of monkeys first inoculated with syphilis and subse- 

 quently treated with subcutaneous inoculations of syphilitic material. He always 

 clarifies the serums and extracts at as short an interval as possible before perform- 

 ing the experiment. Meier finds also that the serum rapidly -deteriorates and 

 either loses its deflecting power, or, more usually, causes deflection even with 

 extracts of normal organs. 



There are various precautions to be observed in regard to both antigen and 

 antibody. Wassermann and his colleagues(2) make the following observations : 



1. Many extracts and serums of themselves alone pick up complement, the 

 amount of extract or serum required varying with the individual preparation and 

 in the case of each, varying with age; 0.1 cubic centimeter of a serum may not 

 block when it is fresh, but after standing it will do so. 



2. Any extract or serum showing even the slightest trace of cloudiness or 

 precipitation will deflect and is unfit for use until centrifugated. 



3. Frequently a serum or extract which is perfectly clear, absorbs complement. 

 This is especially true of those which have stood long before being clarified by 

 centrifugation or before inactivation. 



To avoid this non-specific deflection of complement centrifugate and inactivate 

 immediately after obtaining the serums and extracts ; preserve carefully under 

 aseptic precautions in completely filled tubes, closed by fusing the glass, and 

 keep at a constant freezing temperature away from light and air; centrifugate 

 as frequently as is necessary; determine in each individual experiment the non- 

 deflecting dose, and never use a large one. 



