R. L. KAHN 851 



former extraction period the antigen shows a tendency to throw down a precipitate 

 on standing. An extraction period of three days at about 21° C. was chosen as a 

 standard. Regarding the amount of cholesterol in the antigen, experiments indicated 

 that 0.6 per cent gave optimum results with sera. 



PRODUCING AN UNSTABLE ANTIGEN SUSPENSION 



In addition to the proper concentration of antigenic lipoids in a given antigen, the 

 instability of the antigen-saline suspension is of the utmost importance. The sus- 

 pension employed in the Kahn test contains comparatively large lipoid particles of 

 such instability that they disperse as soon as they come in contact with serum. If the 

 serum is from a syphilitic patient, precipitates appear in a few minutes; if from a non- 

 syphilitic individual, no precipitate appears. 



The following experiment illustrates the antigen-saline relationship. To i-cc. 

 amounts of antigen in a series of tubes are added 0.5, 0.7, o.g, i.i, and 1.3 cc. of 

 normal saline, respectively. On mixing the contents of each tube it will be found that 

 all tubes contain coarse lipoid particles, but of unequal power of dispersion. Thus, the 

 particles in the first tube containing the smallest amount of saline are non-dispersive 

 in additional saline. With the increase in salt solution in the other tubes there is a 

 proportional increase in the dispersive power of the particles. If the i cc. of antigen 

 plus I.I cc. of saline mixture contains particles which are completely dispersed in 

 additional saline, then the particles formed in antigen-saline suspensions in which the 

 amount of saline is greater than i.i cc. are also dispersed on addition of saline. When 

 I cc. of antigen is mixed with as much as 3 cc. of saline, the mixture is usually opales- 

 cent and free from visible particles. 



When employed with serum, the criterion for the antigen-saline suspension is to 

 add to I cc. of antigen the minimum amount of saline which will produce coarse 

 lipoid particles which are completely dispersed in additional saline. These particles 

 will also be dispersed in serum and thus will assure clear-cut negative reactions. In 

 syphilitic serum, how^ever, as already indicated, a specific precipitate will appear 

 within a few minutes. 



The use of a minimum amount of saline in preparing the antigen suspension is 

 closely related to sensitivity with serum. Increasing the saline amount will, in prac- 

 tically all cases, reduce the sensitivity of the antigen suspension. The only time when 

 the criterion for employing the minimum saline amount in the preparation of the 

 antigen suspension is not adhered to is when the suspension is of greater sensitivity 

 than the standard requirement. 



STANDARDIZATION OF ANTIGEN^ 



It was essential to establish a standard for antigen sensitivity because experi- 

 ments have shown that different lots of beef heart may produce antigens of different 

 reactivity. Such a standard was ultimately established as a result of serological and 

 clinical studies. Not a maximum but a conservative degree of antigen sensitivity 

 was chosen and the aim of standardization was to assure uniformity in this character- 

 istic. 



' For complete discussion, see Kahn, R. L., Nagle, N., and Kendrick, P. L.: "Studies on Antigen 

 for the Kahn Test: I. Uniformity in Sensitivity of Standard Antigen," 7. /«/ed. Dis., 41, iii. 1927. 



