METHODS OF TESTING ANTAGONISTIC ACTION 61 



broth and placed in a flask containing the same kind of broth. The 

 test organism is inoculated into the medium inside the sac, and the 

 antagonist into the flask (303). 



Solid Media 



Solid media have also been used extensively for testing the action of 

 antagonists. These media offer certain advantages over liquid media. 

 The following methods are most commonly used: 



Simultaneous inoculation of antagonist and test organism. This method, 

 introduced by Garre (315) in 1887, consists in streaking the an- 

 tagonist and the test organism on the surface of a solidified agar or 

 gelatin medium. The streaks are alternate and may be parallel, radi- 

 ating from a common center, or intersecting at right angles (Fig- 

 ure 4). If the active substance produced by the antagonist does not 

 diffuse for any considerable distance into the medium, the method is 

 not satisfactory. Frost (303) modified this method by inoculating 

 the whole medium with the test organism and, when the medium 

 had hardened, streaking the antagonist across the surface. The first 

 of these came to be known as the anaxogramic method ; the second 

 is often spoken of as the implantation method. The spotting of the 

 two organisms on the plate is illustrated in Figure 5. 



Successive inoculation of the test organism, after the antagonist has al- 

 ready made some growth, so as to enable the active substance to dif- 

 fuse. 



Double plate methods (303). A Petri dish is divided into two parts by 

 means of a small glass tube or rod. After sterilization, one tube of 

 molten agar is heavily inoculated with the antagonist and poured 

 into one half of the plate. When the agar has hardened, another tube 

 of sterile agar is poured into the other half of the plate. Both sides are 

 then streaked with the test organism, each side being equally inocu- 

 lated by separate streaking. This can be done by using a loop bent at 

 nearly right angles; the charged loop is moved from the circumfer- 

 ence toward the glass rod. The loop is then sterilized, recharged with 

 the test culture, and the streak continued on the other side of the 

 plate. The inoculation with the test organism may be made soon 



( after the plate is poured, or the antagonist may be given an opportu- 

 nity to develop before the test organism is streaked thus making the 



