The Chiefs announced Sunday that they had removed Mathieu's injury designation, clearing him to play.
Mathieu suffered a concussion in Kansas City's win over the Buffalo Bills last week, but he had no setbacks in the concussion protocol this week.
Mathieu, who was picked to play in the Pro Bowl, led the Chiefs in interceptions with three this season and returned one for a touchdown.
With Mathieu in the fold, Kansas City has made the past two Super Bowls, winning one, and now has an opportunity for a third straight appearance.
ESPN's Jeremy Fowler contributed to this report.