2. Heat over a Bunsen flame and allow the acid to boil gently for twenty minutes.

NOTE.--A few pieces of pipe-clay or pumice may be placed in the beaker to prevent the "spurting" of the chromic acid.

3. Turn the cover-slips out into a flat gla.s.s dish and wash in running water under the tap until all trace of yellow colour has disappeared.

During the washing keep the cover-slips in motion by imparting a rotatory movement to the dish.

4. Wash in distilled water in a similar manner.

5. Wash in rectified spirit.

6. Transfer the cover-slips, by means of a pair of clean forceps, previously heated in the Bunsen flame to destroy any trace of grease, to a small beaker of absolute alcohol.

Drain off the alcohol and transfer the cover-slips, by means of the forceps, to a wide-mouthed gla.s.s pot, containing absolute alcohol, in which they are to be stored, and stopper tightly.

NOTE.--After once being placed in the chromic acid, the cover-slips must on no account be touched by the fingers.

~Used Slides and Cover-slips.~--Used slides with the mounted cover-slip preparations, and cover-slips used for hanging-drop mounts, should, when discarded, be thrown into a pot containing a 2 per cent. solution of lysol.

After immersion therein for a week or so, even the cover-slips mounted with Canada balsam can be readily detached from their slides.

_Slides._--

1. Wash the slides thoroughly in running water.

2. Boil the slides in water to which "sapon" has been added, for half an hour.

3. Rinse thoroughly in cold water.

4. Dry and polish with a dry cloth.

_Cover-slips._--

1. Wash the cover-slips thoroughly in running water.

2. Boil the cover-slips in 10 per cent. solution of chromic acid, as for new cover-slips.

3. Wash thoroughly in running water.

4. Pick out those cover-slips which show much adherent dirty matter, and rub them between thumb and forefinger under the water tap. The dirt usually rubs off easily, as it has become friable from contact with the chromic acid.

5. Return all the cover-slips to the beaker, fill in _fresh_ chromic acid solution, and treat as new cover-slips.

NOTE.--_Test-tubes, plates, capsules_, etc., which, from long use, have become scratched and hazy, or which cannot be cleaned in any other way, may be dealt with by immersing them in an enamelled iron bath, containing water acidulated to 1 per cent. with hydrofluoric acid, for ten minutes, rinsing thoroughly in water, drying, and polishing.

PLUGGING TEST-TUBES AND FLASKS.

Before sterilisation all test-tubes and flasks must be carefully plugged with cotton-wool, and for this purpose best absorbent cotton-wool (preferably that put up in cylindrical one-pound packets and interleaved with tissue paper--known as surgeons' wool) should be employed.

1. For a test-tube or a small flask, tear a strip of cotton-wool some 10 cm. long by 2 cm. wide from the roll.

2. Turn in the ends neatly and roll the strip of wool lightly between the thumb and fingers of both hands to form a long cylinder.

3. Double this at the centre and introduce the now rounded end into the open mouth of the tube or flask.

4. Now, whilst supporting the wool between the thumb and fingers of the right hand, rotate the test-tube between those of the left, and gradually screw the plug of wool into its mouth for a distance of about 2.5 cm., leaving about the same length of wool projecting.

[Ill.u.s.tration: FIG 24..--Plugging test-tubes: a, cylinder of wool being rolled; b, cylinder of wool being doubled; c, cylinder of wool being inserted in tube.]

The plug must be firm and fit the tube or flask fairly tightly, sufficiently tightly in fact to bear the weight of the gla.s.s plus the amount of medium the vessel is intended to contain, but not so tightly as to prevent it from being easily removed by a s.c.r.e.w.i.n.g motion when grasped between the fourth, or third and fourth, fingers, and the palm of the hand.

For a large flask a similar but larger strip of wool must be taken; the method of making and inserting the plug is identical.

III. METHODS OF STERILISATION.

STERILISING AGENTS.

Sterilisation--i. e., the removal or the destruction of germ life--may be effected by the use of various agents. As applied to the practical requirements of the bacteriological laboratory, many of these agents, such as electricity, sunlight, etc., are of little value, others are limited in their applications; others again are so well suited to particular purposes that their use is almost entirely restricted to such.

The sterilising agents in common use are:

~Chemical Reagents.~--_Disinfectants_ (for the disinfection of gla.s.s and metal apparatus and of morbid tissues).

~Physical Agents.~ HEAT.--(a) _Dry Heat:_

1. Naked flame (for the sterilisation of platinum needles, etc.).

2. m.u.f.fle furnace (for the sterilisation of filter candles, and for the destruction of morbid tissues).

3. Hot air (for the sterilisation of all gla.s.sware and of metal apparatus).

(b) _Moist Heat:_

1. Water at 56 C. (for the sterilisation of certain alb.u.minous fluids).

2. Water at 100 C. (for the sterilisation of surgical instruments, rubber tubing, and stoppers, etc.).

3. Streaming steam at 100 C. (for the sterilisation of media).

4. Superheated steam at 115 C. or 120 C. (for the disinfection of contaminated articles and the destruction of old cultivations of bacteria).

FILTRATION.--