Hamilton graduated from Columbia College in 1827 and was admitted to the New York bar in 1830, but practiced only briefly. He served as commissioner of deeds for the city and county of New York from 1832 through 1833. He was an unsuccessful candidate for New York State Assembly in 1834, and married his wife, Julia Kean (a descendant of a New Yorker who was a New Jersey governor, William Livingston) in 1836.

As a member of the Whig party, Fish was elected to the House of Representatives, defeating Democrat John McKeon and serving in the 28th Congress from New York's 6th District between 1843 and 1845. After losing the election for the 29th Congress, he return to private practice as a lawyer. He was the Whig candidate for lieutenant governor of New York in 1846, but was defeated by Democrat Addison Gardner, in part because of Fish's opposition to the Anti-Rent faction. However, in 1847 Gardner was appointed a judge of the New York Court of Appeals, Fish was elected (November 1847) to complete the term (to January 1849).

He was elected as governor of New York in 1849, defeating John A. Dix and Reuben H. Walworth, and served from November 7, 1848 through November 5, 1850.

He was elected to the United States Senate in March 4, 1851 to replace Daniel S. Dickinson and there he served on the U.S. Senate Committee on Foreign Relations until the end of his term March 3, 1857. He was a Republican for the latter part of his term and was part of a moderately anti-slavery faction. He opposed the repeal of the Missouri Compromise. At the expiration of his term, he traveled with his family to Europe and remained there until shortly before the opening of the American Civil War, when he returned to begin actively campaigning for the election of Abraham Lincoln.

In 1861 and 1862 he was associated with John A. Dix, William M. Evarts, William E. Dodge, A.T. Stewart, John Jacob Astor and other New York men on the Union Defence Committee, which (from April 22, 1861 to April 30, 1862) cooperated with the New York City government in the raising and equipping troops, and disbursed more than $1 million for the relief of New York volunteers and their families.

He also served as Secretary of State between March 17, 1869 to March 12, 1877 under Ulysses S. Grant. He was Grant's longest-serving Cabinet officer.

He conducted the negotiations with Great Britain which resulted in the Treaty of Washington of the May 8, 1871, under which the Alabama claims and the San Juan Boundary Dispute, concerning the Oregon boundary line, were referred to arbitration. He also negotiated the reciprocity treaty of 1875 with the Kingdom of Hawaii.

In 1871 Fish presided at the peace conference at Washington between Spain and the allied republics of Peru, Chile, Ecuador and Bolivia, which resulted in the formulation (April 12) of a general truce between those countries, to last indefinitely and not to be broken by any one of them without three years notice given through the United States.

It was chiefly due to his restraint and moderation that a satisfactory settlement of the Virginius Affair was reached by the United States and Spain in 1873.