The Refuge attracts a great many animals, both resident and migratory. In late August, spectacular concentrations of Franklin gulls gather in the evenings, while mid-September through late October witnesses thousands of migrating sandhill and occasional endangered whooping cranes roosting on the large, flat lake bed. Late October, depending on water conditions, also marks the peak of waterfowl populations with the arrival of up to 25,000 ducks and 20,000 Canada, snow, blue, and white-fronted geese. During the spring and fall, bald eagles are often spotted as they follow migrating waterfowl.

Northern pintail, blue-winged teal, gadwall, and mallards are the Refuge's principal nesting ducks, followed by American wigeon, green-winged teal, northern shoveler, redhead, canvasback, and ruddy ducks.

Other nesting species found on the Refuge include American bittern, piping and upland plovers, and killdeer. Spotted sandpiper, willet, marbled godwit, American avocet, and Wilson's phalarope usually nest on lowlands adjacent to dikes and marsh areas, while ring-necked pheasant, gray partridge, and sharp-tailed grouse are common in suitable upland habitat. Two species of great interest to bird watchers, the Baird's and sharp-tailed sparrow, can also be found.

Because Long Lake National Wildlife Refuge is managed for wildlife diversity in addition to its primary objectives of botulism control and waterfowl production, the Refuge also provides lake, marsh, and upland habitat for such species as white-tailed deer, coyote, fox, raccoon, striped skunk, white-tailed jackrabbit, cottontail rabbit, muskrat, mink, beaver, and badger.

About 500 upland acres are cultivated to provide food and nesting habitat for migratory birds and resident wildlife. Wheat, corn, millet, and sunflowers are planted for food, while stands of mixed sweet clover, alfalfa, and wheat grasses provide nesting habitat. Native and tame grass sites are periodically grazed or hayed to rejuvenate vegetative cover. Local farmers and ranchers assist with upland management in these cooperative programs. Controlled burning is also used to enhance marsh and upland habitat productivity.