Joo Sang-wook made his small screen debut in the 1997 SBS series Docu X. In 2010, he played one of the main characters in the SBS period drama Giant, which won him the New Star Award at the SBS Drama Awards. In 2011, he acted alongside Han Hye-jin and Kim Min-jung in the KBS2 melodrama Thorn Birds, which earned him an Excellence Award nomination at the KBS Drama Awards. In 2012, he became a regular cast member of the KBS2 reality-variety show Qualifications of Men, which scored him the Best Male Newcomer in a Variety Show citation at the KBS Entertainment Awards. In 2013, he starred in the top-rating KBS2 medical drama Good Doctor, which gave him an Excellence Award at the KBS Drama Awards. In 2014, he was paired with Han Ye-seul in the SBS romantic comedy Birth of a Beauty.

Joo Sang-wook and actress Cha Ye-ryun tied the knot in May 2017. They met on the set of the 2015 MBC drama Dazzling Temptation, where they played a couple. Their first child was born in July 2018.