idental contamination.

~Inferior Lethal Coefficient.~--

_Apparatus Required:_

Highly concentrated solutions of the disinfectants.

Sterile test-tubes in which to make dilutions from the
concentrated solutions of the disinfectants.

Hanging-drop slides.

Cover-slips.

Erlenmeyer flask containing 100 c.c. sterile distilled
water.

Case of sterile pipettes, 10 c.c. (in tenths of a cubic
centimetre).

Case of sterile pipettes, 1 c.c. (in tenths of a cubic
centimetre).

METHOD.--

1. Prepare a surface cultivation of the "test" organism B. anthracis
upon nutrient agar in a culture bottle and incubate under optimum
conditions for twenty-four hours; then examine the cultivation
microscopically to determine the absence of spores.

2. Prepare solutions of different percentages of each disinfectant.

3. Make a series of hanging-drop preparations from the agar culture,
using a loopful of disinfectant solution of the different percentages to
prepare the emulsion on each cover-slip.

4. Examine microscopically and note the strongest solution which does
not cause plasmolysis and the weakest solution which does plasmolyse the
organism.

5. Make control preparations of these two solutions and determine the
percentage to be tested.

6. Pipette 10 c.c. sterile water into the culture bottle and suspend the
entire surface growth in it.

7. Transfer the suspension to the Erlenmeyer flask and mix it with the
90 c.c. of sterile water remaining in the flask.

8. Pipette 10 c.c. of the diluted suspension into each of ten sterile
test-tubes.

9. Label one of the tubes "Control" and place it in the incubator at
18 deg. C.

10. Add to each of the remaining tubes a sufficient quantity[11] of a
concentrated solution of the disinfectant to produce the percentage
previously determined upon (_vide_ step 5).

11. Incubate the tubes at 18 deg. C. to 20 deg. C.

12. At hourly intervals remove the control tube and one of the tubes
with added disinfectant from the incubator.

13. Make a subcultivation from both the control and the test suspension,
upon the surface of nutrient agar; incubate under optimum conditions.

14. Observe these culture tubes from day to day until the completion of
seven days, and determine the shortest exposure necessary to cause the
death of vegetative forms.

~Superior Lethal Coefficient.~--

1. Prepare surface cultivations of the