118 



BACTERIOLOGICAL ANALYSIS 



LESSON 



Each time the flask is opened hydrogen must be 

 passed through it again. 



(c) Fractional separation of tetanus bacilli. 



(1) Introduce small quantities of the earth impregnated 

 with tetanus bacilli into three tubes containing liquid grape- 

 sugar agar-agar, and also into three tubes containing liquid 

 grape-sugar gelatine, kept at 60 C. 



(2) Heat them for half an hour at 60 C., and then allow 

 them to cool. 



(3) Now place them in a flask, which must be exhausted 

 in the manner already described (vide p. 114). 



(4) Place flask and tubes in the warm incubator for 

 twenty-four hours. 



(5) Now heat the tubes again at 60 C. for half an hour 

 to one hour ; and place them in a flask, exhausted of air, in 



the warm, incubator. 



(6) Examine the tubes on the 

 second or third day by means of 

 films ; and if the cultures appear to 

 be pure, make subcultures in grape- 

 sugar agar-agar, and in grape-sugar 

 gelatine (deep stab cultures). 



These must be placed in a 

 flask, which then must be 

 exhausted and placed in the 

 warm or cool incubator, as 

 the case may be. 



If the cultures, however, are not, or appear not to be, 

 pure, the tubes should be heated once more at 60 C. for 



FIG. 13. 



a, Thermometer ; fc, thermo- 

 regulator. 



