The western terminus of SR 26 is at
US 19/
98/
27 Alternate (unsigned
SR 55) in
Fanning Springs, near the
Gilchrist/
Levy county line. However the beginning of its run is a short curve towards the north with a connecting
County Road 55A leading to southbound US 19/98/Alt-27, and only begins to move it its signed cardinal direction in
Wilcox east of
the junction between three segments of the
Nature Coast State Trail. Beginning in Wilcox, the route runs parallel to the trail, and continues to do so through
Lottieville, where it carries County Road 341 in a short overlap. The route proceeds east through
Trenton where it is named West Wade Street in the vicinity of the Trenton Elementary School, which is also where the trail beings to move away from the road. West Wade Street becomes East Wade Street at the intersection with
US 129, then the route passes into
Alachua County and
Newberry as West Newberry Road, where it briefly runs through the
city's historic district. After running through the historic district, it intersects with
US 27-
41 (unsigned
SR 45), and continues to maintain the same street name even as it passes through the communities of Jonesville and Tioga before encountering the interchange with
Interstate 75 just west of
Gainesville. campus, approaching
US 441 (13th Street)/
SR 24. East of the interstate, it runs straight east until the intersection with Northwest 57th Street where it curves to the southeast. SR 26 maintains the same street name until the road approaches
State Road 26A, where West Newberry follows the same trajectory onto Southwest Second Avenue, while SR 26 moves onto University Avenue. East of President's Park, the route runs it runs along the northern edge of the
University of Florida campus alongside
Ben Hill Griffin Stadium, followed by a merge with
SR 24 at
U.S. Route 441 (13th Street) and
SR 20 at former
SR 329 (Main Street), separating again near the eastern city limits. Skirting the western shores of
Newnan's Lake as well as the
state forest named for the lake, it approaches the northern shores of the lake entering the Newnans Lake Conservation Area, where it makes a sharp right turn at the eastern terminus of
SR 222. Leaving the preserve area and passing through farmland, the road later crosses
US 301 in
Orange Heights, although a realignment of SR 26 south of the existing intersection was built by the Florida Department of Transportation between 2010 and 2011 which includes an interchange. The next town along the route is
Melrose, where the road enters the
community's historic district, crosses the Alachua-
Putnam County line and intersects
SR 21, before it curves to the northeast and ends at
SR 100 in
Putnam Hall. ==Major intersections==