The earliest predecessor of the Atlantic Coast Line Railroad was the Petersburg Railroad between Petersburg, Virginia and a point near Weldon, North Carolina. A route between Richmond, Virginia and Petersburg was built by the Richmond & Petersburg Railroad, which was founded in 1836. Four years later, the Wilmington and Weldon Railroad, at the time known as the Wilmington and Raleigh and renamed in 1855, completed a route between Weldon and Wilmington, North Carolina. From Wilmington, the Wilmington and Manchester Railroad began operations in 1853 to Florence, South Carolina, where the Northeastern Railroad operated to Charleston, South Carolina.
In 1871, the W&W and the W&M (renamed the Wilmington, Columbia & Augusta) began using the Atlantic Coast Line name to advertise the two lines. An investor from Baltimore, William T. Walters, gained control of these separate railroads after the Civil War, and operated them as a network of independent companies. In 1897–98, most of the South Carolina lines in Walters' system were consolidated under the name of the Atlantic Coast Line Railroad Company of South Carolina. In 1898, as the companies moved towards combining themselves into a single system, the lines in Virginia were combined into the new Atlantic Coast Line Railroad Company of Virginia, and the lines in North Carolina underwent a similar process in 1899, becoming the Atlantic Coast Line Railroad Company of North Carolina.
Around the same time, due to a regulatory climate in Virginia that was better suited to the company than that in other states, the ACL of Virginia took control of the other lines and subsequently shortened its name to the Atlantic Coast Line Railroad Company.
In 1898 the Petersburg Railroad and the Richmond & Petersburg Railroad formally merged, and two years later the combined company took control of the ACL's routes south of Virginia as well as the Norfolk & Carolina Railroad, which operated from Norfolk, Virginia to Tarboro, North Carolina. These mergers created an ACL system reaching from southern Virginia to South Carolina and Georgia.
Other small acquisitions took place in 1901, and in 1902 the ACL took over the Plant System, which operated numerous lines within Florida and Georgia. This same year the ACL took control of the Louisville and Nashville Railroad as well as the Nashville, Chattanooga and St. Louis Railway, though the two were never merged into the ACL and were operated independently. The ACL acquired the East Carolina Railway in 1935, running south from Tarboro to Hookerton, although the 12-mile extension to Hookerton was abandoned in 1933. The ACL's last major acquisition was the Atlanta, Birmingham and Coast Railroad, which it purchased in 1927, though the AB&C was not merged into the ACL until 1945.
By the early 20th century, the railroad had largely reached its final configuration and began to focus on upgrading its physical plant. Later, the railroad's main line from Richmond, Virginia to Jacksonville, Florida had been double-tracked, which benefited the railroad when Florida boomed.
In 1928 the ACL completed a line between Perry, Florida and Drifton, near Monticello, Florida, the last link of the new "Perry Cut-off". This created a more direct route between Chicago and Florida's west coast, one which passed through Macon, Albany, and Thomasville, the route followed by ACL's passenger train The Southland from December 1928 to 1957 when it was rerouted to Jacksonville.
During the Great Depression ACL's freight traffic declined by around 60%, but the railroad survived without declaring bankruptcy; its success in this regard has been attributed to its leadership and careful financial practices, as well as owning the Louisville and Nashville, which remained strong through the Depression.
During World War II ACL's passenger traffic increased 200% and freight traffic 150%. The railroad provided a submarine-proof alternative to coastal shipping, and it also served the fast-emerging military industry in the Southeast. In 1942 Champion McDowell Davis (nicknamed "Champ") became president of the ACL and immediately began an improvement program that included the rebuilding of several hundred miles of track, the installation of modern signalling systems and improvements to freight yards. The railroad spent at least $268 million in upgrading its physical plant during this period.
In 1956 the railroad moved its headquarters from Wilmington, North Carolina to Jacksonville, Florida. Jacksonville was selected from three candidate cities, the other two being Savannah, Georgia and Charleston, South Carolina. Construction of the new office complex was finished four years later, with the move from Wilmington completed over the following weeks.
As early as October 1958 the ACL and competitor Seaboard Air Line Railroad had discussed the possibility of a merger, initiating extensive studies on the potential unified system. The results showed that the merger could save considerable money through savings incurred and reduced expenditures to the amount of $38 million annually. Two years later, the merger was approved by shareholders of both railroads. In 1963, a merger between the two companies was approved by the Interstate Commerce Commission, however, petitions for reconsideration were filed leading to a court decision to remand the approval of the merger on May 13, 1965, citing the Clayton Antitrust Act. Following another round of court decisions in 1966, the merger was allowed to proceed, and did so on July 1, 1967. The result was the creation of the Seaboard Coast Line.