'3$ METHODS OF CULTIVATION OF BACTERIA. 



The gelatin and agar media (vide infra} are treated in the 

 same way. 



" i (b). Glucose Broth. To the other constituents of i (a) there 

 is added i or 2 per cent of grape sugar. The steps in the prep- 

 aration are the same. Glucose being a reducing agent, no free 

 oxygen can exist in a medium containing it, and therefore glu- 

 cose broth is used as a culture fluid for anaerobic organisms. 



1 (c\ Glycerin Broth. The initial steps are the same as in 

 I (a\ but after filtration 6 to 8 per cent of glycerin (sp. grav. 

 1.25) is added. This medium is especially used for growing the 

 tubercle bacillus when the soluble products of the growth of the 

 latter are required. 



2. Agar Media (French, " gelose " ). The disadvantage of 

 gelatin is that at the blood temperature (38 C.), at which most 

 pathogenic organisms grow best, it is liquid. To get a medium 

 which will be solid at this temperature, agar is used as the stiffen- 

 ing agent instead of gelatin. Unlike the latter, which is a proteid, 

 agar is a carbohydrate. It is derived from the stems of various 

 seaweeds growing in the Chinese seas, popularly classed together 

 as " Ceylon Moss." The best for bacteriological purposes is 

 that consisting of the thin dried stem of the seaweed itself. 



2 (a). Ordinary Agar. To preserve as far as possible the 

 relative proportions of the ingredients throughout the process 

 to the end, it will be found most advantageous to ascertain the 

 weight of the finished product just previous to filtration, for 

 then one can accurately make up deficiencies due to evapora- 

 tion, or adjust excesses by further boiling down. To this end 

 a large saucepan is taken and its weight ascertained, and, it 

 being required to make 1000 grammes of agar, 1500 c.c. of water 

 are poured into the pan and set over the flame of a triple Bunsen 

 burner, or a laboratory furnace, and brought to the boil. At this 

 juncture 1 5 grammes of the dry agar are shredded up and dropped 

 in, along with 2.5 grammes of Liebig's meat extract. The boiling 

 is continued until the agar is quite dissolved, usually occupying 

 about thirty minutes, when the scum which rises to the surface 

 should be skimmed off, the fire lowered until boiling ceases, and 

 then 10 grammes of peptone and 5 grammes of common salt should 

 be gradually dusted into the fluid with constant stirring to pre- 

 vent the formation of large lumps of peptone, the pan replaced 

 over the flame and contents boiled until all the peptone is com- 



