40 



SOLID MEDIA 



Warm the mixture at a gentle heat in an enamelled saucepan, stirring 

 constantly to prevent the gelatin sticking to the bottom. When the gelatin 

 is dissolved, boil for two or three minutes. 



5. The medium is now very acid ; add soda solution carefully, testing the 

 reaction with litmus paper after each addition. The end reaction should be 

 neutral or very slightly alkaline. 



6. Autoclave for 5 minutes at 115 C. in a flask or enamelled vessel, to 

 precipitate earthy phosphates. 



7. On taking out of the autoclave pour the hot fluid on to a moistened 

 Chardin paper fixed in a hot water funnel : the filtration must be done in 



the warm, otherwise the gelatin will solidify before it 

 has filtered. 



A more simple method consists in filtering through a 

 glass funnel fitted into a flat- bottomed flask, the flask and 

 funnel being placed in the autoclave [or steamer], which is 

 heated to 100 C. Filtration is quite easy under these con- 

 ditions. 



Hot water funnel. This piece of apparatus consists of a 

 copper funnel mounted on legs (fig. 32), and lined by a second 

 glass funnel, the delivery tube of which, passing through 

 the neck of the metal funnel, is made to fit it closely with an 

 india-rubber bung. Pour water into the space between the 

 two funnels through a small lateral opening provided for the 

 purpose, and heat the apparatus by means of a Bunsen 

 burner placed beneath a side tube projecting from the lower 

 part of the metal funnel. The temperature should not be 

 raised to boiling point, otherwise the water will be driven 

 out through the inlet tube. Heat the funnel before pouring 

 the gelatin on it. Several patterns of this apparatus are 

 made ; one useful form is that in which the metal funnel has 

 two metal walls, and the glass funnel fits inside the inner wall. 



8. Collect the filtered liquid in a flask and tube it at once before it has 

 had time to solidify in quantities of 10-15 c.c. in each tube. 



A small glass funnel should always be used for tubing to avoid soiling the mouth 

 of the tubes, as has been already explained (p. 32). 

 The medium should be perfectly clear. 



9. Plug the tubes : sterilize at 110 C. for 20 minutes, taking care that 

 the temperature does not reach 115 C. [Sterilization may be effected 

 equally by heating in the steamer to 100 C. for 20 minutes on each of three 

 successive days.] 



Notes. (a) Gelatin prepared in this way is quite clear and absolutely trans- 

 parent. If the liquid be slightly cloudy after filtering, add the white of an egg 

 beaten up in 50 to 100 c.c. of water, mix thoroughly and autoclave for 5 minutes 

 at 115 C. ; then after filtering through Chardin paper the mixture is perfectly 

 clear. It is advised that this method be employed as seldom as possible as it is 

 not without influence on the melting point of the gelatin. 



(6) The low temperature at which gelatin melts (23- 25 C.) being a disadvantage 

 in the use of the medium, bacteriologists have attempted to raise the melting point 

 by modifying the method of preparation. Many of these modifications seem to 

 be of no practical value : the author has never found any advantage in using car- 

 bonate of soda for neutralization as recommended by Bertarelli, and similarly 

 Roux's method of sterilization at 100 C. on several successive occasions appears 

 uselessly to complicate the preparation of the medium without offering any cor- 

 responding advantage. 



By following the instructions given above, and provided that (1) a good quality 

 of gelatin be used, (2) neutralization be stopped at the neutral point or when the 

 liquid is very slightly alkaline, and (3) a temperature of 115 C. be never exceeded, 

 a 10 per cent, gelatin is easily obtained which does not melt below 25 C. ; and 



FIG. 32. Hot water 

 funnel. 



