22
A . M O S C O N A , O . A . T R O W E L L A N D E . N . W l L L M E R
solution of known composition, which may be further supplemented
at will with definite constituents, offers many advantages, and it is
the use and construction of such artificial media that is described in
more detail in Chapter 3. There are, however, several points in connexion with the preparation and storage of media in general, which are of
fundamental importance for most forms of culture, and which will
be considered briefly here.
A. G L A S S W A R E
A vital necessity for all forms of Tissue Culture is extreme cleanliness
of all the glassware used not only for culture vessels, but also for storage
and transference of media, for handling tissues or for any other purposes
connected either with the medium or the tissues. Detergents are on the
whole not to be trusted; they are very difficult to remove from glass
surfaces and are often toxic. Chromic acid mixtures are contra-indicated
because the chromate may dissolve in the glass and then slowly come
out into the culture fluids. The safest way is to wash the glassware in a
solution of pure soap in very hot water, rinsing off under the hot tap.
If the water is hard, sodium bicarbonate and tribasic sodium phosphate
can be added as softeners. After washing, the glassware should then
be immersed in approximately normal nitric acid or hydrochloric acid
for a short time, after which it is thoroughly rinsed in several changes
of distilled water and dried in the oven. Provided that pure soap is
used and the rinsing is thorough, none of these chemicals cause any
apparent harm.
B. W A T E R
The water used for making media and for the final washing of glassware must be very pure. It should be distilled at least twice, the last
distillation being carried out in an all-glass apparatus provided with
an efficient spray-trap. De-ionization by ion-exchange resins is inadequate, as this does not remove organic contaminants such as
bacterial toxins and other biologically active compounds; it may in
fact add them—bacteria have been known to grow on the resin column.
Furthermore, a certain amount of resin dissolves in the water. It is
advisable to test the final product for traces of heavy metals by the very
sensitive dithizone colour reaction (Sandell, 1950).
C. OSMOTIC PRESSURE
The osmotic pressure of the medium must be adapted to that of the
tissues under investigation. This is, of course, well recognized in relation
to the tissues of different groups of animals whose body-fluids differ
A . M O S C O N A , O . A . T R O W E L L A N D E . N . W l L L M E R
solution of known composition, which may be further supplemented
at will with definite constituents, offers many advantages, and it is
the use and construction of such artificial media that is described in
more detail in Chapter 3. There are, however, several points in connexion with the preparation and storage of media in general, which are of
fundamental importance for most forms of culture, and which will
be considered briefly here.
A. G L A S S W A R E
A vital necessity for all forms of Tissue Culture is extreme cleanliness
of all the glassware used not only for culture vessels, but also for storage
and transference of media, for handling tissues or for any other purposes
connected either with the medium or the tissues. Detergents are on the
whole not to be trusted; they are very difficult to remove from glass
surfaces and are often toxic. Chromic acid mixtures are contra-indicated
because the chromate may dissolve in the glass and then slowly come
out into the culture fluids. The safest way is to wash the glassware in a
solution of pure soap in very hot water, rinsing off under the hot tap.
If the water is hard, sodium bicarbonate and tribasic sodium phosphate
can be added as softeners. After washing, the glassware should then
be immersed in approximately normal nitric acid or hydrochloric acid
for a short time, after which it is thoroughly rinsed in several changes
of distilled water and dried in the oven. Provided that pure soap is
used and the rinsing is thorough, none of these chemicals cause any
apparent harm.
B. W A T E R
The water used for making media and for the final washing of glassware must be very pure. It should be distilled at least twice, the last
distillation being carried out in an all-glass apparatus provided with
an efficient spray-trap. De-ionization by ion-exchange resins is inadequate, as this does not remove organic contaminants such as
bacterial toxins and other biologically active compounds; it may in
fact add them—bacteria have been known to grow on the resin column.
Furthermore, a certain amount of resin dissolves in the water. It is
advisable to test the final product for traces of heavy metals by the very
sensitive dithizone colour reaction (Sandell, 1950).
C. OSMOTIC PRESSURE
The osmotic pressure of the medium must be adapted to that of the
tissues under investigation. This is, of course, well recognized in relation
to the tissues of different groups of animals whose body-fluids differ