He was the founding editor of the Archives of Pathology and Laboratory Medicine in 1926 and edited several other medical journals.

At the age of 14, Hektoen enrolled in Luther College in Iowa, during which time he developed a friendship with Johan K. Schreiner, a Norwegian physician who inspired him to pursue medicine.

[3]: 1–5 After graduating from Luther College with a Bachelor of Arts degree in 1883, Hektoen spent the next year taking pre-medical courses at the University of Wisconsin.

To finance his medical education, Hektoen worked as an attendant and later a druggist in the Northern Hospital for the Insane in Oshkosh, Wisconsin.

[3]: 6  During his internship Hektoen trained under Christian Fenger, who influenced his decision to specialize in pathology.

In 1918 he became a member of the National Academy of Sciences, and he received, among other honors, the Order of St. Olaf in 1929 and the Distinguished Service Medal of the American Medical Association in 1942.

[2]: 167  In 1894, he published The technique of post-mortem examination, a textbook on autopsies for medical students at the Cook County Hospital.

[1][7][8] In 1917, while studying immunity in rabbits, he published a description of what is now known as the anamnestic response;[2]: 169–70 [4] and in 1933, working with William H. Welker, he introduced the use of aluminum hydroxide as an adjuvant to increase the immunogenic potential of injected antigens.