~Sugar Agar.~--

1. Prepare nutrient agar (_vide_ page 167, sections 1 to 8). Measure out 1000 c.c.

2. Weigh out glucose, 20 grammes (= 2 per cent.), and dissolve in the clear agar.

3. Tube, and sterilise as for nutrient agar.

NOTE.--Other "sugar" media are prepared by subst.i.tuting a corresponding percentage of lactose, maltose (or any other of the substances referred to under "Sugar Media," page 177) for the glucose.

~Iron Bouillon.~--

1. Measure out nutrient bouillon, 1000 c.c. (_vide_ page 141, sections 1 to 6).

2. Weigh out ferric tartrate, 1 gramme (= 0.1 per cent.), and dissolve it in the bouillon.

3. Tube, and sterilise as for bouillon.

NOTE.--The lactate of iron may be subst.i.tuted for the tartrate.

~Lead Bouillon.~--

1. Measure out nutrient bouillon, 1000 c.c. (_vide_ page 163, sections 1 to 6).

2. Weigh out lead acetate, 1 gramme (= 0.1 per cent.), and dissolve it in the bouillon.

3. Tube, and sterilise as for bouillon.

~Nitrate Bouillon.~--

1. Measure out nutrient bouillon, 1000 c.c. (_vide_ page 163, sections 1 to 6).

2. Weigh out pota.s.sium nitrate, 5 grammes (= 0.5 per cent.), and dissolve it in the bouillon.

3. Tube, and sterilise as for bouillon.

NOTE.--The nitrate of sodium or ammonium may be subst.i.tuted for that of pota.s.sium, or the salt may be added in the proportion of from 0.1 to 1 per cent. to meet special requirements.

~Iron Peptone Solution (Pakes).~--

1. Weigh out peptone, 30 grammes, and emulsify it with 200 c.c. tap water, previously heated to about 60C.

2. Wash the emulsion into a litre flask with 800 c.c. tap water.

3. Weigh out salt, 5 grammes, and sodium phosphate, 3 grammes, and dissolve in the mixture in the flask.

4. Heat the mixture in the steamer at 100 C. for thirty minutes, to complete the solution of the peptone, and filter into a clean flask.

5. Fill into tubes in quant.i.ties of 10 c.c. each.

6. Add to each tube 0.1 c.c. of a 2 per cent. neutral solution of ferric tartrate. (A yellowish-white precipitate forms.)

7. Sterilise as for nutrient bouillon.

~Lead Peptone Solution.~--

Prepare as for iron peptone solution but in step 6 subst.i.tute 0.1 c.c.

of a 1 per cent. neutral aqueous solution of lead acetate.

~Nitrate Peptone Solution (Pakes).~--

1. Weigh out Witte's peptone, 10 grammes, and emulsify it with 200 c.c.

ammonia-free distilled water previously heated to 60C.

2. Wash the emulsion into a flask and make up to 1000 c.c., with ammonia-free distilled water.

3. Heat in the steamer at 100 C. for twenty minutes.

4. Weigh out sodium nitrate, 1 gramme, and dissolve in the contents of the flask.

5. Filter through Swedish filter paper.

6. Tube, and sterilise as for nutrient bouillon.

~Litmus Bouillon.~--

1. Measure out nutrient bouillon, 1000 c.c. (_vide_ page 163, sections 1 to 6).

2. Add sufficient sterile litmus solution to tint the medium a dark lavender colour. (Media rendered +10 will usually react very faintly alkaline or occasionally neutral to litmus.)

3. Tube, and sterilise as for bouillon.

~Rosolic Acid Peptone Solution.~--

1. Weigh out rosolic acid (corallin), 0.5 gramme, and dissolve it in 80 per cent. alcohol, 100 c.c. Keep this as a stock solution.

2. Measure out peptone water (Dunham), 100 c.c., and rosolic acid solution, 2 c.c., and mix.

3. Heat in the steamer at 100 C. for thirty minutes.

4. Filter through Swedish filter paper.