3. Neutralise to litmus by the cautious addition of citric acid solution 4 per cent. (Do not neutralise with _mineral_ acid.)

4. Heat in the steamer at 100 C. for one hour to coagulate all the proteid.

(If the whey is cloudy when removed from the steamer allow it to stand for forty-eight hours in the ice chest and then decant off the clear fluid--or filter through a Berkefeld filter candle.)

5. Filter into a sterile flask.

6. Tint the whey with litmus solution to a deep purple red.

7. Tube, and sterilise as for milk.

~Litmus Whey (Petruschky).~--

1. Measure out into a flask

Fresh milk 1000 c.c.

2. Add

Hydrochloric acid (or glacial acetic acid) 1.5 c.c.

and boil.

3. Filter off coagulated casein.

4. Neutralise to litmus by the addition of n/1 caustic soda solution and boil. Whey now cloudy and acid again.

5. Again neutralise to litmus by addition of n/10 caustic soda solution.

6. Filter.

7. Tint the whey with neutral litmus solution to a deep purple colour.

8. Tube and sterilise as for milk.

~Litmus Whey Gelatine.~--

1. Measure out milk 1000 c.c. into a tared 2-litre flask.

2. Add hydrochloric acid (or glacial acetic acid) 1.5 c.c. and boil for five minutes.

3. Filter off the casein, and make the whey faintly alkaline to litmus.

4. Weigh out

Peptone 10 grammes

and emulsify in a few cubic centimeters of the whey and return to the flask.

5. Weigh out

Gelatine 50 grammes

add it to the whey in the flask and incorporate the mixture by bubbling through live steam.

6. Clear with egg and filter.

7. Make the weight of the medium ma.s.s to the calculated figure for one litre (1060 grammes) by the addition of distilled water.

8. Weigh out

Dextrose 15 grammes

and dissolve in the fluid whey gelatine.

9. Add sterile litmus solution to the required tint.

10. Tube and sterilise for twenty minutes in steamer at 100C. on each of five successive days.

This medium will remain semi-fluid at the room temperature, and may be used for cultures in the cool or hot incubator.

~Litmus Whey Agar~ is prepared in a similar manner to Whey Gelatine, with the subst.i.tution of 15 grammes of agar for the gelatine.

~Malt Extract Solution (Hersch.e.l.l).~--

1. Measure into a flask distilled water 1000 c.c.

2. Weigh out

Extractum malti (malt extract) 25 grammes

and add to distilled water in flask.

3. Boil for five minutes, allow to stand, and decant off clear fluid from sediment.

4. Tube and sterilise as for nutrient bouillon.

_Media for the Study of Earth Bacteria, Nitrogen Fixers._

~Earthy Salts Agar (Lipman and Brown).~--(_For the enumeration of soil organisms._)

1. Measure out