70 METHODS OF CULTIVATION OF BACTERIA. 



they are sterilised in the steam steriliser (p. 38). After 

 sterilisation the gelatine is melted and one tube inoculated 

 with the mixture containing the anaerobes ; the second is 

 inoculated from the first, and the third from the second, 

 as in making ordinary gelatine plates. After inoculation 

 the gelatine is kept liquid by the lower ends of the tubes 

 being placed in water at about 30 C., and hydrogen is 

 passed in through tube x for twenty 

 minutes. The gas-supply tubes are 

 then completely sealed off at x and 

 /, and each test-tube is rolled as in 

 Esmarch's method till the gelatine 

 solidifies as a thin layer on the 

 internal surface. A little hard 

 paraffin may be run between the 

 rim of the test-tube and the stop- 

 per, and round the perforations for 

 the gas-supply tubes, to ensure that 

 the apparatus is air-tight. The 

 gelatine is thus in an atmosphere 

 of hydrogen in which the colonies 

 ma Y develop. The latter may be 

 examined and isolated in a way 

 which will be presently described. The method is ad- 

 mirably suited for all anaerobes which grow at the ordinary 

 temperature. 



(b] In glucose agar (Vignal's method). Three pieces 

 of ordinary quill-tubing (bore about J inch) about a foot 

 long are taken, the ends are tapered in the gas flame, and 

 they are then sterilised. Three ordinary test-tubes, a, b, c, 

 of glucose agar are now boiled in a beaker, cooled to 42 

 C, and inoculated with the bacterial mixture as for plate 

 cultures, i.e., b from a and c from b. A current of hydrogen 

 being passed through the pieces of quill-tubing to expel the 

 air, the contents of each of the test-tubes are now sucked 

 up with the mouth into one of the quill-tubes and both the 

 ends of the latter sealed off in the flame. The agar will 

 solidify in situ and the tubes may be incubated. The 



taining anaerobes. 



