BACTERICIDAL METHODS 127 



a number of small test-tubes sterilised and plugged with cotton- 

 wool. We can then make any required dilution of a young 

 bacterial culture in bouillon as follows : To each of a number 

 of tubes we add '9 c.c. of '8 per cent, solution of sodium chloride. 

 To the first tube (a) we add '1 c.c. of the bacterial culture, and 

 thoroughly shake up the mixture ; to the second (6) we add 

 1 c.c. of the contents of (a), and shake up ; to the third tube 

 (c) we add '1 c.c. of the contents of (6), and so on. It is thus 

 evident that '1 c.c. of the contents of (a) will correspond to 

 '01 c.c., and '1 c.c. of (b) to '001 c.c. of the original culture ; any 

 minimi fraction can thus be readily obtained. In the making 

 of all mixtures of serum and bacteria it is essential that none of 

 the latter shall escape the action of the former, e.y. by remaining 

 on a part of the mixing vessel with which the serum does not 

 come in contact. 



(a) Method of Neisser and Wechsbery. A series of small 

 plugged sterile tubes is taken, and to each we add '5 c.c. of 

 8 per cent, sodium chloride solution, and a given quantity, say 

 5^5. c.c., of a young bouillon culture to be tested. To the 

 several tubes in series we then add varying amounts of the 

 fresh serum whose action is to be observed, e.y. '2 c.c., '1 c.c., 

 05 c.c., '025 c.c., etc. The contents of each tube are then 

 made up to 1 c.c. with salt solution, and a few drops of sterile 

 bouillon are added to each tube. The tubes are then well shaken 

 and placed in the incubator at 37 C. for three hours, to allow 

 the serum to act. (Of course several series of such tubes may be 

 prepared and placed in the incubator for varying periods of time ; 

 we can thus observe when the bactericidal effect reaches the 

 maximum.) At the end of the given period of time a small 

 quantity, say '05 c.c., of the contents of each tube is added to a 

 tube of melted agar (cooled to about 40 C.) ; each agar tube is 

 then shaken, and the contents are poured out into a sterile Petri 

 capsule. The other tubes are similarly treated, and the Petri 

 capsules are placed in the incubator for a suitable period of time. 

 The number of colonies in each can then be noted. Of 

 course gelatine can be substituted for the agar in the plates if 

 desired. 



(b) Wright's Method. A twenty-four hours 1 bouillon culture is 

 used, and various dilutions with sterile bouillon are made according 

 to the method described on p. 58 : thus 5-, 10-, 20-, 50-, 100-, 

 1000-, etc., fold dilutions may be prepared. A small quantity, 

 .say 1 c.mm., of the fresh serum to be tested is mixed with an 

 equal amount of the bacterial culture, and the mixture is placed 

 in a small capillary tube which is sealed at the ends ; similar 



