o act for a few minutes, then wash off the
excess in running water.

The penetrating power of stains is increased by (a) physical
means--e. g., heating the stain; (b) chemical means--e. g., by the
addition of carbolic acid, 5 per cent. aqueous solution; caustic
alkalies, 2 per cent. aqueous solutions; water saturated with aniline
oil; borax, 0.5 per cent. aqueous solution.

The most commonly used dyes for cover-slip film preparations are the
aniline dyes.

(A) Basic:
(a) Methylene-blue.
(b) Gentian violet.
(c) Fuchsin.

These dyes are kept in saturated alcoholic (90 per cent.) solutions so
that decomposition may be retarded.

Two or three drops of alcoholic solution of these dyes to, say, 4 c.c.
water, usually makes a sufficiently strong staining fluid for cover-slip
film preparations.

Carbolic methylene-blue (C.M.B.) and carbol fuchsin (C.F.) are prepared
by covering the cover-slip with 5 per cent. solution of carbolic acid
and adding a few drops of the saturated alcoholic solution of
methylene-blue or fuchsin respectively to it. For aniline gentian violet
(A.G.V.) the stain is added to a saturated solution of aniline oil in
water.

(d) Thionine blue.
(e) Bismarck brown.
(f) Neutral red.
(B) Acid:
(a) Eosin, aqueous yellowish.
(b) Safranine.

These dyes are kept in 1 per cent. aqueous solution to which is added 5
per cent. of alcohol, as a preservative. They are generally used in this
form.

A few nuclear stains (carmine, haematoxylin) are occasionally used more
especially in "section" work.

_Decolourisation._--After overstaining, films may be decolourised by
washing for a longer or shorter time in one of the following reagents
arranged in ascending order of power

1. Water.
2. Chloroform.
3. Acetic acid, 1 per cent.
4. Alcohol.
5. Alcohol absolute, } equal parts.
Acetic acid, 1 per cent., }

{Hydrochloric, 1 per cent. aqueous solution.
{Hydrochloric, 1 per cent. Alcoholic
{ (90 per cent.) solution.
6. Mineral acids: {Sulphuric, 25 per cent. aqueous solution.
{Nitric, 33 per cent. aqueous solution.

_Counterstaining._--Use colours which will contrast with the first
stain; e. g.,

Vesuvin, }
Neutral red, }for films stained by methylene-blue or
Eosin,