2 . M E T H O D S
29
size of the fragment and the ease with which it can be penetrated.
It also depends upon the extent of the differences between the internal
and external fluids. Thus the conditions necessary for the culture of
small pieces of tissue and of isolated cells are in this way much more
exacting than those required for larger pieces of tissue, and the behaviour of the cells in each case may be different.
Most studies so far made on the survival of cells and tissues in culture
indicate that cells may tolerate abnormal conditions for a time, but
the further the conditions are removed from the normal, the shorter is
survival likely to be. Cells and tissues inevitably lose important constituents to inadequate media and attention is called to the chapter
on synthetic media (p. 124) and on amino acid metabolism (p. 285)
where some of these problems are discussed in more concrete form.
Another factor in determining the survival-time of cultures in simple
media is the extent to which the cells can utilize or are damaged by
the products of the breakdown of other tissues and cells which die and
autolyse in the culture.
I I . T I S S U E C U L T U R E
Although it would seem logical to begin with a description of cell
culture and then proceed via tissue culture to organ culture, historically it was tissue culture that developed first. Tissue culture paved the
way for the later development of cell and organ culture, so it will be
best to deal with it first.
A. H A N G I N G - D R O P M E T H O D
Of the various tissue culture techniques commonly used, the hangingdrop method was the earliest to yield successful results and we shall
therefore begin with its description. The method, in its simplest form,
will be described in some detail since many of its features are of importance in relation to tissue culture as a whole.
1. Fluid Medium
A small piece (about 1 mm
3
) of tissue is cut out as cleanly and aseptically as possible with cataract knives, or other instruments appropriate
for the tissue in question, and with the minimum amount of tearing or
crushing. The tissue is meanwhile kept moist with a suitable medium.
It is placed on a rigorously cleaned and sterilized coverslip made from a
reasonably insoluble and non-toxic glass, or from quartz or mica. It is
then covered with just sufficient protective or nutritive fluid to keep it
29
size of the fragment and the ease with which it can be penetrated.
It also depends upon the extent of the differences between the internal
and external fluids. Thus the conditions necessary for the culture of
small pieces of tissue and of isolated cells are in this way much more
exacting than those required for larger pieces of tissue, and the behaviour of the cells in each case may be different.
Most studies so far made on the survival of cells and tissues in culture
indicate that cells may tolerate abnormal conditions for a time, but
the further the conditions are removed from the normal, the shorter is
survival likely to be. Cells and tissues inevitably lose important constituents to inadequate media and attention is called to the chapter
on synthetic media (p. 124) and on amino acid metabolism (p. 285)
where some of these problems are discussed in more concrete form.
Another factor in determining the survival-time of cultures in simple
media is the extent to which the cells can utilize or are damaged by
the products of the breakdown of other tissues and cells which die and
autolyse in the culture.
I I . T I S S U E C U L T U R E
Although it would seem logical to begin with a description of cell
culture and then proceed via tissue culture to organ culture, historically it was tissue culture that developed first. Tissue culture paved the
way for the later development of cell and organ culture, so it will be
best to deal with it first.
A. H A N G I N G - D R O P M E T H O D
Of the various tissue culture techniques commonly used, the hangingdrop method was the earliest to yield successful results and we shall
therefore begin with its description. The method, in its simplest form,
will be described in some detail since many of its features are of importance in relation to tissue culture as a whole.
1. Fluid Medium
A small piece (about 1 mm
3
) of tissue is cut out as cleanly and aseptically as possible with cataract knives, or other instruments appropriate
for the tissue in question, and with the minimum amount of tearing or
crushing. The tissue is meanwhile kept moist with a suitable medium.
It is placed on a rigorously cleaned and sterilized coverslip made from a
reasonably insoluble and non-toxic glass, or from quartz or mica. It is
then covered with just sufficient protective or nutritive fluid to keep it