Ann Bancroft grew up in St. Paul, Minnesota. She described her family as one of risk takers. It is reported that she struggled with a learning disability, but nevertheless graduated from St. Paul Academy and Summit School. Bancroft was a camper and staff member at YMCA Camp Widjiwagan. Bancroft became a wilderness instructor and a gym teacher in Minneapolis (at Clara Barton Open School) and St. Paul.

She gave up her teaching post in 1986 in order to participate with the"Will Steger International North Pole Expedition". She arrived at the North Pole together with five other team members after 56 days using dogsleds. This made Bancroft the first woman to reach the North Pole on foot and by sled.

She was also the first woman to ski across Greenland. In 1992 and 1993 Bancroft led a four woman expedition to the South Pole on skis. This expedition was the first group of women to reach the South Pole on skis. Her achievements led to her induction into the National Women's Hall of Fame for the United States.

She currently co-owns an exploration company, Bancroft Arnesen Explore, with Liv Arnesen. In March 2007, Bancroft and Arnesen were taking part in a trek across the Arctic Ocean to draw attention to the problem of global warming. However, according to The Washington Post, the expedition was called off "after Arnesen suffered frostbite in three of her toes, and extreme cold temperatures drained the batteries in some of their electronic equipment."

Bancroft also received a number of other awards and honors. She lives in Minnesota near the St. Croix River. Bancroft is an out lesbian and in 2006, she publicly campaigned against a proposed amendment to the Minnesota Constitution to prohibit any legal recognition of marriages or civil unions between members of the same sex.