Carrel was a young surgeon in 1894, when the French President Sadi Carnot was assassinated with a knife. Carnot bled to death due to severing of his portal vein, and Surgeons who treated the President felt that the vein could not be successfully reconnected. This left a deep impression on Carrel, and he set about developing new techniques for suturing blood vessels. The technique of "triangulation", using three stay-sutures as traction points in order to minimize damage to the vascular wall during suturing, was inspired by sewing lessons he took from an embroideress and is still used today. Julius Comroe wrote: "Between 1901 and 1910, Alexis Carrel, using experimental animals, performed every feat and developed every technique known to vascular surgery today." He had great success in reconnecting arteries and veins, and performing surgical grafts, and this led to his Nobel Prize in 1912.

In 1902 Alexis Carrel went from being a skeptic of the visions and miracles reported at Lourdes to being a believer in spiritual cures after experiencing a healing of Marie Bailly that he could not explain. The Catholic journal Le nouvelliste reported that she named him as the prime witness of her cure. Alexis Carrel refused to discount a supernatural explanation and steadfastly reiterated his beliefs, even writing a book describing his experience, though it was not published until four years after his death. This was a detriment to his career and reputation among his fellow doctors, and feeling he had no Future in academic Medicine in France, he emigrated to Canada with the intention of farming and raising cattle. After a brief period, he accepted an appointment at the University of Chicago and two years later at the Rockefeller Institute of Medical Research.

In 1906 he joined the newly formed Rockefeller Institute of Medical Research in New York where he spent the rest of his career. There he did significant work on tissue cultures with pathologist Montrose Thomas Burrows. In the 1930s, Carrel and Charles Lindbergh became close friends not only because of the years they worked together but also because they shared personal, political, and social views. Lindbergh initially sought out Carrel to see if his sister-in-law's heart, damaged by rheumatic fever, could be repaired. When Lindbergh saw the crudeness of Carrel's machinery, he offered to build new equipment for the scientist. Eventually they built the first perfusion pump, an invention instrumental to the development of organ transplantation and open heart surgery. Lindbergh considered Carrel his closest friend, and said he would preserve and promote Carrel's ideals after his death.

Carrel was also interested in the phenomenon of senescence, or aging. He claimed that all cells continued to grow indefinitely, and this became a dominant view in the early 20th century. Carrel started an experiment on January 17, 1912, where he placed tissue cultured from an embryonic chicken heart in a stoppered Pyrex flask of his own design. He maintained the living culture for over 20 years with regular supplies of nutrient. This was longer than a chicken's normal lifespan. The experiment, which was conducted at the Rockefeller Institute for Medical Research, attracted considerable popular and scientific attention.

According to Gwen Terrenoire, writing in Eugenics in France (1913-1941) : a review of research findings, "The foundation was a pluridisciplinary centre that employed around 300 researchers (mainly statisticians, Psychologists, physicians) from the summer of 1942 to the end of the autumn of 1944. After the liberation of Paris, Carrel was suspended by the Minister of Health; he died in November 1944, but the Foundation itself was "purged", only to reappear in a short time as the Institut national d’études démographiques (INED) that is still active." Although Carrel himself was dead most members of his team did move to the INED, which was led by famous demographist Alfred Sauvy, who coined the expression "Third World". Others joined Robert Debré's "Institut national d'hygiène" (National Hygiene Institute), which later became the INSERM.

During World War I (1914–1918), Carrel and the English Chemist Henry Drysdale Dakin developed the Carrel–Dakin method of treating wounds based on chlorine (Dakin's solution) which, preceding the development of antibiotics, was a major medical advance in the care of traumatic wounds. For this, Carrel was awarded the Légion d'honneur.

Carrel advocated the use of gasses to rid humanity of "defectives", thus endorsing the scientific racism discourse. His endorsement of this idea began in the mid-1930s, prior to the Nazi implementation of such practices in Germany. In the 1936 German introduction of his book, at the publisher's request, he added the following praise of the Nazi regime which did not appear in the editions in other languages:

In 1935, Carrel published a book titled L'Homme, cet inconnu (Man, The Unknown), which became a best-seller. In the book, he attempted to outline a comprehensive account what is known and more importantly unknown of the human body and human life "in light of discoveries in biology, physics, and medicine", to elucidate problems of the modern world, and to provide possible routes to a better life for human beings.

In 1937, Carrel joined Jean Coutrot’s Centre d’Etudes des Problèmes Humains - Coutrot’s aim was to develop what he called an "economic humanism" through "collective thinking." In 1941, through connections to the cabinet of Vichy France President Philippe Pétain (specifically, French industrial Physicians André Gros and Jacques Ménétrier) he went on to advocate for the creation of the Fondation Française pour l’Etude des Problèmes Humains (French Foundation for the Study of Human Problems) which was created by decree of the Vichy regime in 1941, and where he served as 'regent'.

Carrel co-authored a book with famed pilot Charles A. Lindbergh, The Culture of Organs, and worked with Lindbergh in the mid-1930s to create the "perfusion pump," which allowed living organs to exist outside the body during surgery. The advance is said to have been a crucial step in the development of open-heart surgery and organ transplants, and to have laid the groundwork for the artificial heart, which became a reality decades later. Some critics of Lindbergh claimed that Carrel overstated Lindbergh's role to gain media attention, but other sources say Lindbergh played an important role in developing the device. Both Lindbergh and Carrel appeared on the cover of Time magazine on June 13, 1938.

In his later life he returned to his Catholic roots. In 1939 he met with Trappist monk Alexis Presse on a recommendation. Although Carrel was skeptical about meeting with a priest, Presse ended up having a profound influence on the rest of Carrel's life. In 1942, he said "I believe in the existence of God, in the immortality of the soul, in Revelation and in all the Catholic Church teaches." He summoned Presse to administer the Catholic Sacraments on his death bed in November 1944.

The Foundation made many positive accomplishments during its time. Yet it was also behind the origin of the December 16, 1942 Act inventing the "prenuptial certificate", which had to precede any marriage and was supposed, after a biological examination, to insure the "good health" of the spouses, in particular in regard to sexually transmitted diseases (STD) and "life hygiene" (sic). The institute also conceived the "scholar book" ("livret scolaire"), which could be used to record students' grades in the French secondary schools, and thus classify and select them according to scholastic performance.

The foundation was at the origin of the October 11, 1946, law, enacted by the Provisional Government of the French Republic (GPRF), which institutionalized the field of occupational Medicine. It worked on demographics (Robert Gessain, Paul Vincent, Jean Bourgeois-Pichat), on economics, (François Perroux), on nutrition (Jean Sutter), on habitation (Jean Merlet) and on the first opinion polls (Jean Stoetzel). "The foundation was chartered as a public institution under the joint supervision of the ministries of Finance and public health. It was given financial autonomy and a budget of forty million francs—roughly one franc per inhabitant—a true luxury considering the burdens imposed by the German Occupation on the nation’s resources. By way of comparison, the whole Centre National de la Recherche Scientifique (CNRS) was given a budget of fifty million francs."

Carrel's experiment by some was never successfully replicated, and in the 1960s Leonard Hayflick and Paul Moorhead proposed that differentiated cells can undergo only a limited number of divisions before dying. This is known as the Hayflick limit, and is now a pillar of biology.

In 1972, the Swedish Post Office honored Carrel with a stamp that was part of its Nobel stamp series. In 1979, the lunar crater Carrel was named after him as a tribute to his scientific breakthroughs.

In February 2002, as part of celebrations of the 100th anniversary of Charles Lindbergh's birth, the Medical University of South Carolina at Charleston established the Lindbergh-Carrel Prize, given to major contributors to "development of perfusion and bioreactor technologies for organ preservation and growth". Michael DeBakey and nine other Scientists received the prize, a bronze statuette [2] created for the event by the Italian Artist C. Zoli and named "Elisabeth" after Elisabeth Morrow, sister of Lindbergh's wife Anne Morrow, who died from heart disease. It was in fact Lindbergh's disappointment that contemporary medical Technology could not provide an artificial heart pump which would allow for heart surgery on her that led to Lindbergh's first contact with Carrel.

Experiments with the Carrel-Dakin method were depicted in the 2014 BBC miniseries, The Crimson Field.

L. Hayflick has shown that a cell has a limited number of divisions, equal to the so called "Hayflick’s Limit." However, L. Franks and others (Loo et al. 1987; Nooden and Tompson 1995; Frolkis 1988a), have shown that the number of cell divisions can be considerably greater than that stipulated by the "Hayflick Limit", having practically no limit at all.