)[5] In 1875, the Kingston Road Tramway operated a horsecar service along what would become today's Queen Street East.

[11] In 1957, to accommodate construction of the Gardiner Expressway, the streetcar tracks were removed from Lake Shore Boulevard between Roncesvalles Avenue and the Humber River, and the bridge crossing the rail corridor was demolished.

TTC CEO Andy Byford wanted a car-free corridor along one of the city's two busiest streetcar lines (either 501 Queen or 504 King) to prevent bunching, gaps and service delays.

Byford would ban general automobile traffic along the streetcar corridor but allow cyclists, taxicabs and delivery trucks.

On January 3, 2016, the 501 Queen route was split into two sections at Humber Loop in order to make its service more reliable.

[16] In January 2017, construction began for approximately 15 months on various projects west of Roncesvalles Avenue, including rebuilding the bridge over the Humber River carrying streetcar tracks.

[17] Other work included replacing track and overhead along portions of the Queensway and Lake Shore Boulevard as well as at the Humber Loop.

[23] From May to September 2017, all streetcar service along the entire 501 Queen route was replaced by buses to permit multiple construction projects along Queen Street to take place, including the replacement of an overhead pedestrian walkway at the Eaton Centre, which required overhead wires to be removed.

[17] Since March 31, 2021, the 501 Queen route has been partially closed due to construction activity, including track replacement and the extension of the exclusive streetcar right-of-way on the Queensway west of Roncesvalles Avenue.

The TTC had to address rush-hour crowding due to the accelerated retirement of the ALRV fleet, the last of which made their final voyages on September 2, 2019.

[43][44] By 2007, critics of the TTC's management of this line argue that small delays at one end ripple into 30-40 minute waits at the other.

Transit proponents such as Steve Munro have long claimed that Route 501 would be better off if it were split into two or three overlapping segments.

[45] In late 2009, the TTC conducted an experiment of splitting the 501 streetcar route into two overlapping segments, as recommended by critics to alleviate bunching, gaps and short turns when delays occurred.

It was determined that splitting the route increased short turns by 90%, required more streetcars and resulted in poorer service.

To address these problems and make service more predictable for riders, the TTC again split the 501 Queen route.

Transfer-free service through Humber Loop was still provided by 3 runs in the morning, in the late evening and overnight.

[16][52][53] The split was suspended starting from January 8, 2017,[17] due to construction projects affecting Humber Loop.

[54] From June 21, 2020, most 501 Queen streetcars operated the full route between Neville Park and Long Branch loops.

This ended the split in 501 Queen service at Humber Loop where passengers from Long Branch had to change streetcars to continue downtown.

[55] From Neville Park Loop, the route proceeds westward in mixed traffic on Queen Street East.

West of Humber Loop, the line passes through a short streetcar tunnel under a railway corridor and the Gardiner Expressway.

The line emerges from the tunnel onto Lake Shore Boulevard West and proceeds westward in mixed traffic to Long Branch Loop, where both the 507 and late evening 501 streetcars terminate.

It was created on October 28, 1935, merged into an extended 501 Queen on March 26, 1995[57] and reinstated as a separate route on November 19, 2023.