With the Blue Jays trailing 2-1 in the fourth, Colby Rasmus singled while Kelly Johnson and Travis Snider each drew a walk off Rick Porcello (7-6) to load the bases with two outs. That brought up light-hitting catcher Mathis, who hammered a 1-0 pitch over the glove of Tigers left fielder Quintin Berry, chasing home all three runners.
Anthony Gose followed with a single to drive in Mathis and Snider later added a two-run homer as Toronto went on to take the opener of a three-game series.
Carlos Villanueva (6-0) got the victory, allowing a pair of runs on four hits in a five-inning start.
Porcello (7-6) was touched for five runs on six hits over six frames.
Miguel Cabrera and Prince Fielded socked back-to-back homers in the first inning for second-place Detroit, which came into the contest one-half game behind the American League Central-leading Chicago White Sox.