During the Mexican–American War, he joined Colonel Stephen Watts Kearney's expedition to California and painted accounts of the campaign, as well as aspects of the Oregon Territory.

His appreciation and portrayal of the American West is valued, and today his few surviving works are held by national and numerous regional museums.

He taught himself painting skills, and at the age of 20 moved to Detroit, the largest city in the Michigan Territory, and started doing itinerant work.

[2] Stanley spent four weeks there and worked intensely through the next three months to complete his numerous paintings of individuals and tribal groups.

In early 1846 in Cincinnati, Ohio, he and Dickerman exhibited a gallery of 85 paintings of Indians, which received favorable reviews there and in Louisville, Kentucky.

[1] At the outbreak of the Mexican War in 1846, Stanley was appointed a draftsman for the Corps of Topographical Engineers to Colonel Stephen Watts Kearney's expedition to California and the Oregon Territory.

He traveled further north to the Oregon and Washington territories to paint landscapes and various Native American tribes, and worked through part of 1848.

[1][3] That year Stanley traveled to Hawaii, where he spent nearly twelve months painting portraits of King Kamehameha III, his wife, and the royal family.

[1] In 1852, he gained a major exhibit in Washington, DC, at the Smithsonian Institution of his Native American Gallery, which attracted much attention in the city.

[1] In 1853, Stanley was appointed chief artist at a salary of $125 per month[4] for Isaac I. Stevens' expedition to survey a northern railroad route to the Pacific Coast; he made the most of this chance for travel and work in the Northwest.

Stanley observed gatherings of nearly 1,500 Assiniboine, traveled to a distant Blackfoot (Piegan) village, and saw a large hunting party of several hundred, including families from the Pembina area near the Canada–US border.

Stanley painted and sketched many Northwest landmarks, which were reproduced in lithographs for inclusion in Stevens' last volume of the Pacific Railroad Reports.

The party returned that year to the East, crossing the Isthmus of Panama from the west and arriving by ship in New York in January 1854.

[1] After his return, Stanley worked intensely at painting and organizing a large panorama of western scenes from the northern survey route.

Probably written in the winter of 1868–1869, these include his preface, as well as pages describing three plates: a Plains Indian encampment, Chinook burial ground, and a buffalo hunt.

His surviving works are held by national museums as well as numerous regional institutions: National: Regional: Stanley's art was celebrated in an exhibition entitled "Painted Journeys-The Art of John Mix Stanley" that opened June 2015 at the Buffalo Bill Center of the West in Cody, Wyoming.