Han Jae-seok debuted as an actor in the 1994 MBC series Last Lovers. In 2000, he became part of the main cast of the hit MBC drama All About Eve. In 2002, he acted alongside Kim Hyun-joo and Kim Ji-ho in the 40-episode SBS series Glass Slippers. That same year, he appeared in the SBS drama The Great Ambition, which won him an Excellence Award at the SBS Drama Awards. In 2004, he was cast in the Taiwanese TV series Amor de Tarapaca, opposite Taiwanese actress and singer Ruby Lin. In 2008, he was nominated for an Excellence Award at the KBS Drama Awards for his work in Women in the Sun. In 2010, he earned another Excellence Award nomination at the KBS Drama Awards for the sageuk series Merchant Kim Man-deok. In 2011, he joined the cast of the Chinese action film Speed Angels.

In 2004, Han Jae-seok was embroiled in a draft-dodging scandal along with fellow actors Song Seung-heon and Jang Hyuk. It was revealed that they illegally avoided military service by manipulating their medical examination results. Amid public backlash, they enlisted in the military that same year and were discharged in 2006. In April 2013, he married his Merchant Kim Man-deok co-star Pak Sol-mi. They welcomed their first child on March 23, 2014.