'On the Permanent Life of Tissues outside of the Organism' reports Alexis Carrel's 1912 experiments on the maintenance of tissue in culture media. At the time, Carrel was a French surgeon and biologist working at the Rockefeller Institute in New York City. In his paper, Carrel reported that he had successfully maintained tissue cultures, which derived from connective tissues of developing chicks and other tissue sources, by serially culturing them. Among all the tissue cultures Carrel reported, one was maintained for more than two months, whereas previous efforts had only been able to keep tissues in vitro for three to fifteen days. Carrel’s experiments contributed to the development of long-term tissue culture techniques, which were useful in the study of embryology and eventually became instrumental in stem cell research. Despite later evidence to the contrary, Carrel believed that as long as the tissue culture method was accurately applied, tissues kept outside of the organisms should be able to divide indefinitely and have permanent life.

Boris Ephrussi studied fruit flies, yeast, and mouse genetics and development while working in France and the US during the twentieth century. In yeast, Ephrussi studied how mutations in the cytoplasm persisted across generations. In mice he studied the genetics of hybrids and the development of cancer. Working with George Wells Beadle on the causes of different eye colors in fruit flies, Ephrussi's research helped establish the one-gene-one-enzyme hypothesis. Ephrussi helped create new embryological techniques and contributed the theories of genetics and development.

Alexis Carrel was a doctor and researcher who studied tissue cultures. He continued Ross Granville Harrison's research and produced many improvements in the field of tissue culture and surgery. He was the recipient of the 1912 Nobel Prize in Physiology or Medicine for his development of surgical techniques to repair blood vessels. Carrel was born on 28 June 1873 in Sainte-Foy-les-Lyon, France, to Anne-Marie Ricard and Alexis Carrel Billiard. His father died when he was five years old. Carrel earned a bachelor's degree in letters in 1889 and another in science in 1890 from St. Joseph's Day School in Lyon, France. He entered medical school at the age of seventeen and was regarded as a good but not exceptional student. The assassination of Sadi Carnot, a French politician visiting Lyon who was stabbed in the abdomen and died from the loss of blood, further interested him in surgery.

In an effort to develop tissue culture techniques for long-term tissue cultivation, French surgeon and biologist Alexis Carrel, and his associates, produced and maintained a series of chick heart tissue cultures at the Rockefeller Institute in New York City. From 1912 to 1946, this series of chick heart tissue cultures remained alive and dividing. Since the duration of this culture greatly exceeded the normal chick life span, the cells were deemed immortal. Although this conclusion was challenged by further experiments in the 1960s, the publicity surrounding the immortal chick heart tissue significantly influenced the concept of cell immortality and cellular aging from the 1920s through the 1960s. Carrel's experiment convinced many biologists to accept immortality as an intrinsic property of all cells, not just the cell line through which genetic material is passed to offspring, called the germ line. Consequently, the phenomenon of cellular aging was regarded not as an intrinsic characteristic, but was attributed to external factors such as the accumulation of waste products within the cell.

In "The Outgrowth of the Nerve Fiber as a Mode of Protoplasmic Movement," Ross Granville Harrison explores the growth of nerve fibers in vitro. The purpose of this experiment was to test two possible hypotheses for the growth of nerve fibers. Santiago Ramón y Cajal suggested that nerve growth is due to the extension of nerve fibers as they push through tissue. Victor Hensen's syncytial theory proposed an opposing view of nerve growth. He proposed that each neuron was connected by threads of cytoplasm and the successful connections stimulated further differentiation of the correct neural connections. Using hanging drop tissue cultures, Harrison provided significant evidence for Ramón y Cajal's theory by showing discrete cell membranes between cells and observing the growth of individual neurons.

The HeLa cell line was the first immortal human cell line that George Otto Gey, Margaret Gey, and Mary Kucibek first isolated from Henrietta Lacks and developed at The Johns Hopkins Hospital in Baltimore, Maryland, in 1951. An immortal human cell line is a cluster of cells that continuously multiply on their own outside of the human from which they originated. Scientists use immortal human cell lines in their research to investigate how cells function in humans. Though the HeLa cell line has contributed to many advancements in biomedical research since the twentieth century, its usage in medical research has been controversial because Lacks did not consent to having her cells used for such purposes. As of 2020, scientists continue to use the HeLa cell line for numerous scientific advancements, such as the development of vaccines and the identification of many underlying disease mechanisms.

In the 1910s, Alexis Carrel, a French surgeon and biologist, concluded that cells are intrinsically immortal. His claim was based on chick-heart tissue cultures in his laboratory that seemed to be able to proliferate forever. Carrel's ideas about cellular immortality convinced his many contemporaries that cells could be maintained indefinitely. In the 1960s, however, Carrel's thesis about cell immortality was put into question by the discovery that human diploid cells can only proliferate for a finite period. As it was gradually recognized that chick cells only have a finite proliferative life span in vitro as well, historians and scientists alike attempted to identify experimental errors that could have led to the extremely long life of Carrel's "immortal" chick-heart tissue cultures. Those reassessments not only point out potential experimental mistakes in pioneer tissue culture work in the early twentieth century, but are also relevant to current discussions about the different life spans of germ line cells, embryonic and adult stem cells, normal somatic cells, and cancer cells.

Alexis Carrel, the prominent French surgeon, biologist, and 1912 Nobel Prize laureate for Physiology or Medicine, was one of the pioneers in developing and modifying tissue culture techniques. The publicized work of Carrel and his associates at the Rockefeller Institute established the practice of long-term tissue culture for a wide variety of cells. At the same time, some aspects of their work complicated the operational procedures of tissue culture. Thus Carrel's legacy had a mixed influence on the development of tissue culture techniques, which have been widely used in the fields of embryology and stem cell research.

The hanging drop tissue culture is a technique utilized in embryology and other fields to allow growth that would otherwise be restricted by the flat plane of culture dishes and also to minimize the surface area to volume ratio, slowing evaporation. The classic hanging drop culture is a small drop of liquid, such as plasma or some other media allowing tissue growth, suspended from an inverted watch glass. The hanging drop is then suspended by gravity and surface tension, rather than spreading across a plate. This allows tissues or other cell types to be examined without being squashed against a dish.

Leo Loeb developed an experimental approach to studying cancer and pioneered techniques for tissue culture and in vitro tissue transplantation which impacted early-to-mid twentieth century experimental embryology. Loeb received his medical degree from the University of Zurich in 1897. As part of his doctorate, he completed a thesis on the outcomes of tissue transplantation in guinea pigs. Loeb's thesis inspired a life-long interest in tissue transplantation. His research culminated in greater than 400 publications, including a book called The Biological Basis of Individuality, in which he demonstrated the potential immortality of certain mammalian tissues.