Born in Flehingen, Germany, at age 14 he moved to Rockport, Indiana. Within a couple of years he had enrolled at the Indiana University, where he studied under David Starr Jordan. Eigenmann received a bachelor's degree in 1886, and soon after went to California, where he met Rosa Smith, herself already becoming known for her work on West Coast fisheries. They married on August 20, 1887, and then went to Harvard University, where they studied the collections made by Louis Agassiz and Franz Steindachner, and produced the first of a series of joint publications.
They moved to San Diego, California in 1888, where he worked as curator of a natural history society, and helped found the San Diego Biological Laboratory. He received his PhD from Indiana in 1889, and took up a professorship of zoology there in 1891. In 1892, famed scientist Albert C. L. G. Günther financed Eigenmann's first expedition, a trip throughout western North America where many new species were collected. Subsequent explorations focussed on the blind fishes and salamanders of caves in Indiana, then in the caves of Texas, Missouri, and Cuba.
After a trip to the University of Freiburg in 1906-7, in 1908 Eigenmann became the dean of Indiana's graduate school. In the same year he secured support from the Carnegie Museum for a trip to South America, and in September 1908 the Carnegie British Guiana Expedition got underway. They returned with 25,000 specimens, resulting in the description of 128 new species and 28 new genera. Subsequent trips went to Colombia (1912) and the Andes (1918). Rosa continued to collaborate with him during the Indiana years, but less frequently; of their five children, one daughter was disabled and a son was eventually institutionalized, a burden that was mostly borne by Rosa.
Eigenmann's later years were spent writing up reports on his previous expeditions, and assisting younger colleagues in mounting their own trips. He was elected to the National Academy of Sciences in 1923.
When Carl suffered a stroke in 1927, the family returned to San Diego. He died in April 1927.