Prior to his sophomore season, Castonzo made the move from right tackle to left tackle where he started all 14 games in 2008. Boston College ranked 25th in the country in tackles for a loss allowed (4.77) and 49th in sacks allowed (1.69). Castonzo earned a College Football News All-Sophomore First Team selection.
In 2009, Castonzo was listed at No. 9 on Rivals.com′s preseason offensive tackle power ranking. Following a productive junior season, Castonzo was selected 2009 First Team All-Atlantic Coast Conference as voted on by the 40 members of the Atlantic Coast Sports Media Association. He set the record for career starts with 54 straight at Boston College and was a Rhodes Scholar nominee in 2010.
Castonzo became the Colts starting left offensive tackle for the 2011 season. He started 12 games during his rookie season and the Colts finished 2-14 without longtime starting quarterback Peyton Manning. On April 28, 2014, the Colts picked up the fifth-year, $7.438 million option in his rookie contract. In week 11 of the 2014 season, Castonzo recorded his first career reception and touchdown, catching a 1-yard TD pass from Andrew Luck. He played 1,115 snaps during the 2014 season, the most of any offensive lineman in the NFL.