Tim DeKay was born June 12, 1963 to Jim and Jill DeKay in Lansing, New York, where he and his brother Jamey grew up. Growing up DeKay enjoyed athletics, and he played both varsity basketball and baseball, the latter of which is a generational tradition in his family. He also enjoyed the arts, and performed in his high school's production of Oliver!. He attended Le Moyne College to study Business and Philosophy, where he also played baseball, eventually deciding to pursue a career in theater. After graduating with a bachelor's degree in Business administration in 1985, he planned for a career in Business or law, but eventually realized that the theater was his true calling. After taking a few courses in directing at Syracuse University, he attended Rutgers University, where he received a Master of Fine Arts and met his wife, Actress Elisa Taylor. The couple lives in California with their two children, daughter Danna and son Jamis.

In 2009, DeKay began starring in the USA Network comedy-drama White Collar as FBI agent Peter Burke. He made his directorial debut in the episode "Stealing Home", which premiered in 2012. In March 2015, it was announced that he will play FBI agent Duval Pritchard in the FOX pilot, Frankenstein.

DeKay returned to Le Moyne College in May 2010 as the commencement speaker, and was awarded the degree of Doctor of Humane Letters honoris causa. He is a member of the Actors Studio. He taught as an adjunct professor at the University of Southern California.