Tissue Culture

TISSUE CULTURE. The body is composed of countless myriads of cells assembled in tissues and organs. The cells belong to several types : wandering cells such as blood leucocytes and tissue macrophages which roam through the entire organism; epithelial cells, the noblest elements of the body, which form the glands, brain, skin, etc. ; connective tissue cells present every where under various appearances, etc. They are the units of a strictly organized community. Their existence is bound to that of the whole. If the whole is taken apart, its component tissues and organs die of ter a short time. The cultivation of tissues consists precisely in maintaining in a condition of active life cells that have been removed from the body. The aim of this method is to study the innate properties of each cell type, and the mechanism of their organization into tissues and organs. Its ultimate purpose is the discovery of the laws which express the relations between the tissues and the body fluids, and the building up of cell soci ology.

History.—The method of tissue culture may be considered the indirect outcome of studies made independently by several biolo gists on the survival of cells outside of the organism. However, the first chapter of its history was written in 19°7 by R. G. Harrison in the beautiful experiments where nerve fibres were caused to grow in vitro from fragments of the central nervous system of frog embryos. In 191o, these experiments were extended by other workers to adult and embryo tissues of birds and mammals, which were observed to survive in a drop of plasma. But after a few days, degeneration began and death occurred. In these experi ments, as well as in those of Harrison, the tissues were not culti vated in the proper sense of the word. They were only in a condi tion of survival. A second chapter was begun in 1911 when A. Carrel developed a procedure by which the death of the surviving tissues could be indefinitely postponed and found, in 1912, that embryo proteins possess the power of promoting the unlimited proliferation of tissue cells. Then, it became possible really to cultivate cells, that is, to cause them to manufacture new proto plasm from the substances contained in their medium. A long interruption occurred in the progress of these investigations due to the World War. But, in 1921, the development of procedures per

mitting the main cell types to be obtained in pure cultures opened a third and important period in the history of the method. Blood and tissue macrophages, iris epithelium, cartilage cells, thyroid cells, Malpighian epithelium, lens cells, and several types of malig nant cells were isolated from other tissue elements and maintained in a pure state. At the same time, it became possible to measure accurately the rate of growth of the strains of tissue cells. New procedures were perfected by which living tissues could be main tained indefinitely in flasks where they were washed, fed, and measured without being disturbed and without danger of infection from bacteria. These great practical improvements rendered the method of tissue culture very much easier, and widely extended its application to physiological and pathological problems. Al though it has not as yet reached its complete development, it may be considered the most powerful instrument now possessed for the investigation of cell physiology.

Technique.

The handling of the delicate and minute struc tures which compose the tissues demands elaborate techniques and a long apprenticeship for their mastery. But the principles of these techniques are very simple. The cells are cultivated in flat, round flasks made of pyrex glass and varying in size from 3 to 8cm. in diameter (see fig.). The flasks possess a wide, oblique neck 3cm. in length. Dust and bac teria from the air do not fall into the flask when it is opened, but remain on the wall of the neck where they are destroyed by flaming. By means of a spatula, the tissue cells are conveniently handled inside the flasks. After being flamed, the neck is hermetically sealed with a rubber cap. It is thus possible entirely to protect the tissues against bacteria which are their deadly enemies, and also maintain in the air of the container the proper amount of humidity and of carbon dioxide. Through the bottom of the flask, the tissues may be examined with a microscope at a magnification of about 120 diameters. When the cells must be studied in greater detail, they are removed from the flask with a spatula, placed on a thin cover-glass. and exam ined while still living, at a magnification of 1,5oo diameters.