9. Measure out and add to the medium

Picric acid, 1 per cent. aqueous solution 20 c.c.

10. Tube and sterilise as for nutrient agar.

~China Green Agar (Werbitski).~--

1. Liquefy and measure out into a sterile flask

Nutrient agar 1000 c.c.

2. Adjust the reaction accurately to +13 and filter.

3. Measure out and mix thoroughly with the fluid agar

China green 0.2 per cent. aqueous solution 15 c.c.

4. Tube and sterilise as for nutrient agar.

~Malachite Green Agar (Loeffler).~--

1. Liquefy and measure out into a sterile flask

Nutrient agar 1000 c.c.

2. Weigh out

Dextrose 10 grammes.

and dissolve in nutrient agar.

3. Adjust the reaction to +3, and filter.

4. Measure out and mix thoroughly in the fluid agar

Malachite green, 0.1 per cent. aqueous solution 16 c.c.

for ~"weak"~ medium.

_4a._ To the filtered agar add

Malachite green, 2 per cent. aqueous solution 25 c.c.

for ~"strong"~ medium.

5. Tube and sterilise as for nutrient agar.

~Double Sugar Agar (Russell).~--

1. Liquefy and measure out into a sterile flask

Nutrient agar 1000 c.c.

2. Add 100 c.c. litmus solution to the fluid agar.

3. Weigh out and dissolve in the fluid agar.

Lactose 10 grammes Dextrose 10 grammes.

4. Render the reaction of the medium neutral to litmus paper by the cautious addition of normal caustic soda.

5. Tube in quant.i.ties of 10 c.c. and sterilise in the steamer at 100 C.

for twenty minutes on each of three successive days.

6. Store for use in a cool dark place.

_B. Diphtheriae._

~Glycerine Blood-serum.~--

1. Prepare blood-serum as described, page 168, sections 1 to 4.

2. Add 5 per cent. pure glycerine.

3. Complete as described above for ordinary blood-serum, sections 5 to 7.

NOTE.--Different percentages of glycerine--from 4 per cent.

to 8 per cent.--are used for special purposes. Five per cent. is that usually employed.

~Blood-serum (Loeffler).~--

1. Prepare nutrient bouillon (_vide_ page 163), using meat extract made from veal instead of beef.

2. Add 1 per cent. glucose to the bouillon, and allow it to dissolve completely.

3. Now add 300 c.c. clear blood-serum (_vide_ page 168, sections 1 to 4) to every 100 c.c. of this bouillon.

4. Fill into sterile tubes and complete as for ordinary blood-serum.

~Blood-serum (Lorrain Smith).~--

1. Collect blood-serum (_vide_ page 168, sections 1 to 4), as free from haemoglobin as possible.

2. Weigh out 0.15 per cent. sodium hydrate and dissolve it in the fluid (or add 0.375 c.c. of dekanormal soda solution for every 100 c.c. of serum).

3. Tube, and stiffen at 100 C. in the serum insp.i.s.sator.