Interstate 65 (I-65) in the US state of Indiana traverses from the south-southeastern Falls City area bordering Louisville, Kentucky, through the centrally located capital city of Indianapolis, to the northwestern Calumet Region of the Hoosier State which is part of the Chicago metropolitan area. The Indiana portion of I-65 begins in Jeffersonville after crossing the Ohio River and travels mainly north, passing just west of Columbus prior to reaching the Indianapolis metro area. Upon reaching Indianapolis, the route alignment of I-65 begins to run more to the northwest and subsequently passes Lafayette on that city's east and north sides. Northwest of there, in west-central Jasper County, the route again curves more northward as it approaches the Calumet Region. Shortly after passing a major junction with I-80 and I-94, I-65 reaches its northern national terminus in Gary at I-90 which is carried on the Indiana East–West Toll Road. I-65 covers 261.27 miles (420.47 km) in the state of Indiana. This is one of the principal Interstate Highways that cross the state, and, more specifically, intersect at the city of Indianapolis, that has given the state the nickname of "Crossroads of America".

Jeffersonville is a city and the county seat of Clark County, Indiana, United States, situated along the Ohio River. Locally, the city is often referred to by the abbreviated name Jeff. It lies directly across the Ohio River to the north of Louisville, Kentucky, along I-65. The population was 49,447 at the 2020 census.