In one pan of the trip balance place the counterpoise of the tared flask (or its equivalent in weights) together with the weights making up the _calculated medium weight_. In the opposite pan place the flask containing the medium mass. Now add boiling distilled water from a wash bottle until the two pans are exactly balanced. 7. Titrate and estimate the reaction of the medium mass; control the result. Calculate the amount of soda solution required to make the reaction of the medium mass +10 (i. e., calculate for 1000 c.c., less the quantity used for the titrations). 8. Add the necessary amount of soda solution and heat in the steamer at 100° C. for twenty minutes, to precipitate the phosphates, etc. 9. Allow the medium mass to cool to 60° C. Well whip the whites of two eggs, add to the contents of the flask and replace in the steamer at 100° C. for about half an hour (until the egg-albumen has coagulated and formed large, firm masses floating on and in clear gelatine). 10. Filter through papier Chardin into a sterile flask. 11. Tube in quantities of 10 c.c. 12. Sterilise in the steamer at 100° C. for twenty minutes on each of three consecutive days--i. e., by the discontinuous method. ~Nutrient Agar-agar.~-- 1. Weigh a 2-litre flask and note the weight--or counterpoise exactly. 2. Measure out double strength meat extract, 500 c.c., into the "tared" flask. 3. Weigh out and mix 10 grammes of peptone, 5 grammes of salt, and 20 grammes of powdered agar, and make into a thick paste with 150 c.c. distilled water, and add to the meat extract in the flask; place the flask in a water-bath. 4. Arrange the steam can and water-bath as already directed (for the preparation of gelatine) and figured. 5. Bubble live steam (at 100° C.) through the medium mass, for twenty-five minutes, by which time complete solution of the agar is effected. 6. Now weigh the flask and its contents; then (1035[5] grammes + weight of flask) minus (weight of flask and its contents) equals the weight of water required to make up the bulk of the medium to 1 litre. Add the requisite amount (see preparation of gelatine, page 166, step 6). 7. Titrate, and estimate the reaction of the medium mass; control the result. Calculate the amount of soda solution required to make the reaction of the medium mass + 10 (i. e., calculated for 1000 c.c., less the quantity used for the titrations). 8. Add the necessary amount of soda solution and replace in the steamer for twenty minutes (to complete the precipitation of the phosphates, etc.). 9. Allow the medium mass to cool to 60° C. Well whip the whites of two eggs, add to the contents of the flask, and replace in the steamer at 100° C. for about _one hour_ (until the egg-albumen has coagulated and formed large, firm masses floating on and in clear agar.) 10. Filter through papier Chardin, by the aid of a hot-water funnel, if necessary (Fig. 101), into a sterile flask. 11. Tube in quantities of 10 c.c. or 15 c.c.