[1] The refuge is located between the cities of Buffalo and Rochester and is operated by the United States Fish and Wildlife Service.

The 10,828-acre (43.82 km2) refuge encompasses several large pools (separated by dikes), swamps, meadows, fields, and woodlands.

Kanyoo Trail, located nearby to the east, has two loops (0.33 and 1 mile [0.53 and 1.61 km] long).

The eastern half of the refuge contains a scattering of smaller pools, many marshy areas, fields, and upland forests.

Iroquois National Wildlife Refuge allows hiking, cross-country skiing, snowshoeing, nature observation, and photography.

Prohibited activities include camping, disturbing historical artifacts, swimming, fires, removing wildlife and plants, horseback riding and off-road vehicle use, and carrying firearms except during hunting season (with permit).