After studying law at Toulouse, Lartet – the son of a wealthy landowner – spent the earlier part of his life managing the family estates at St. Guiraud. His interest in paleontology was aroused in the early 1830s when he was shown a locally discovered mastodon tooth. Beginning with his discovery in 1836 of Pliopithecus, the ancestor of the gibbon, he made a number of significant fossil discoveries. In 1856 he found remains of Dryopithecus, the fossil ancestor of the other apes, and in 1868, at Cro-Magnon in the Dordogne, he came across several adult human skeletons, later to be named Cro-Magnon man, that were found to belong neither to Neanderthal nor to modern man. These are the earliest known fossils of humans in Europe. They have modern teeth and jaw bones and a brain capacity averaging about 1300 cubic centimeters.

Lartet's most important work, however, helped to solve one of the major problems of 19th-century science, namely the antiquity of humans. Early proposals that human history could be pushed back into glacial times lacked conclusive proof: attempts to link human remains with the bones of extinct animals were always open to the objection that they were accidental assemblies that had come together at some historic time. Nor was the argument that some of the animal bones had been worked by humans any more persuasive, since the working of ancient bones could have taken place at any time.

In the early 1860s Lartet began a careful study of the caves of the Dordogne and the Pyrenees in collaboration with the British banker Henry Christy. In a cave at La Madelaine in 1863 he found a piece of ivory with a woolly mammoth clearly engraved upon it. Excluding forgery, there seemed no other explanation than that an animal of the ice age and a human witness had coexisted.

Lartet went on to propose a sequence for the Paleolithic based on the presence of animals, but scholars preferred alternative classifications based on tools. The results of his researches were published posthumously in Reliquiae Aquitanicae (1875; Aquitanian Remains), a work that did much to establish the prime importance of the archeological sites of southern France. In 1869 Lartet was appointed professor of paleontology at the museum of the Jardin des Plantes – a post he held until his death.