The refuge is located approximately 70 miles east of Washington, North Carolina. The headquarters entrance road is located off Highway 94, 1.5 miles north of U.S. 264 between Swan Quarter and Engelhard.

Detailed directions to the Mattamuskeet National Wildlife Refuge Visitor Center:

Driving south from Columbia, NC: Start out going south on S Broad St/Alligator River Route/NC-94 toward Branning St. Continue to follow Alligator River Route/NC-94. In 33.4 miles turn left onto Mattamuskeet Refuge Rd. In 1.39 miles, the Visitor Center is on the left.

Driving east from Belhaven, NC: Start out going northwest on W Main St/US-264 Bus W toward Quidley Ln. Take the 3rd right onto US Highway 264 Byp/Pamlico Scenic Byway/US-264 E. Continue to follow Pamlico Scenic Byway/US-264 E. In 33.57 miles turn left onto NC Highway 94/Alligator River Route/NC-94. In 1.5 miles turn right onto Mattamuskeet Refuge Rd. In 1.39 miles Visitor Center is on the left.

Driving west or north starting from Engelhard, NC: Start out going west on US Highway 264/Pamlico Scenic Byway/US-264 W toward Roper Ln. In 11.83 miles turn right onto New Holland Rd. New Holland Rd becomes Mattamuskeet Refuge Rd. Visitor Center is on the left.