He made his professional regular season debut in the Arizona Cardinals' season-opener at the St. Louis Rams and made seven solo tackles and made his first career forced fumble in their 27–24 loss. Mathieu made his first career regular season tackle and forced a fumble on St Louis Rams' tight end Jared Cook. He forced a fumble before Cook crossed into the endzone for a touchdown and it was recovered by Cardinals' linebacker Karlos Dansby for a touchback in the first quarter. The following week, he earned his first career start as a nickelback and made six solo tackles and a pass deflection during a 25–21 victory against the Detroit Lions in Week 2. On September 22, 2013, Mathieu recorded ten combined tackles (nine solo), deflected a pass, and made his first career interception during a 31–7 loss at the New Orleans Saints. Mathieu intercepted a pass by quarterback Drew Brees, that was initially intended for wide receiver Lance Moore, in the third quarter. In Week 4, Mathieu earned his first career start at free safety after Rashad Johnson sustained an injury to his finger while covering a punt the previous week. Mathieu finished the Cardinals' 13–10 win at the Tampa Bay Buccaneers in Week 4 with six solo tackles. On October 6, 2013, he recorded five solo tackles and made his first career sack during the Cardinals' 22–6 win against the Carolina Panthers. Mathieu sacked quarterback Cam Newton for a ten-yard loss in the second quarter. On December 8, 2013, Mathieu made one solo tackle and a pass deflection before exiting the Cardinals' 30–10 victory against the St. Louis Rams with a leg injury. It was later determined Mathieu tore his left ACL and LCL while returning a punt during the third quarter. On December 10, 2013, the Arizona Cardinals placed Mathieu on injured reserve officially ending his rookie season. He completed his rookie season in 2013 with 68 combined tackles (65 solo), nine pass deflections, two interceptions, a sack, and a forced fumble in 13 games and 11 starts. Mathieu was named a first-team All-Pro at the slot cornerback position by Pro Football Focus after the season.