In 2000, he joined Milan’s youth system. He made his first-team debut for the Rossoneri on 28 November 2006, aged only 16, in the Coppa Italia against Brescia, replacing Kakha Kaladze at half-time.

He made his Serie A debut at the age of 17, in Milan’s 3–2 defeat against Udinese on 19 May 2007, when he entered in the 67th minute to replace Giuseppe Favalli. During the 2007–08 season he became the leader and captain of the A.C. Milan Primavera, collecting another appearance for the first-team in the Coppa Italia. The following season, the 19-year-old defender collected an additional three league appearances. He ended his experience at Milan with four appearances in the top flight.

After playing for Italy at under-17 and under-18 levels, he made his debut for the under-19 squad, led by Francesco Rocca, in June 2007. At the 2008 UEFA European U-19 Championship he played in the starting lineup, always in the role of a right full-back. In 2009, he took part in the Mediterranean Games, playing seven games for the under-20 side. On 25 March 2009, he made his debut for the Italy under-21 side in a 2–2 friendly match draw against Austria, which was played in Vienna. From 10 to 12 March 2014, he was called up to the senior national team by coach Cesare Prandelli as part of an internship to evaluate young players ahead of the 2014 FIFA World Cup, and was also part of the next meeting between 14 and 15 April 2014.

On 17 July 2009, Darmian moved to Serie B club Padova on loan until the end of the 2009–10 season. He made his official debut for the club on 28 November 2009, in a home loss against Vicenza. He went on to make 20 appearances and score one goal, as Padova finished the season in 19th place and subsequently won the relegation play-off against Triestina.

On 11 July 2010, it was announced that Matteo Darmian would join Palermo’s pre-season training camp in Austria, after Milan and the Sicilian Serie A club agreed in principle about a transfer move involving him. The move, unveiled as a co-ownership bid, was officially announced by Palermo the next day. The fee was €800,000. Darmian made his debut for the Rosanero on 16 September 2010, in a Europa League game against Sparta Prague, which also earned him his debut in European competition. His Palermo league debut was during the fourth game of the season, coming on for Javier Pastore in a 3–1 win against Juventus. On 6 February 2011, he played his first game as a starter in the league in a 4–2 victory for Palermo away at Lecce, but was substituted in the 33rd minute due to injury. He finished the season with 16 appearances among Serie A, Europa League and Coppa Italia.

After Milan and Palermo renewed the co-ownership on 23 June 2011, he was officially loaned to Serie B side Torino on 12 July 2011. He made his debut for the Granata on 13 August 2011, in the second round of Coppa Italia against Lumezzane; and scored his first goal on 30 October 2011, against Empoli, allowing his team to win 2–1. On 13 November 2013, he was injured against Bari and sidelined for five games. At the end of the season, he was promoted to Serie A, having played 33 games in the league and one in the Coppa Italia.

On 20 June 2012, Palermo redeemed his 50% share from Milan for €500,000 and on 5 July 2012, his return to Torino was formulised under a co-ownership agreement for €825,000. With Torino he played 30 games in Serie A and two in the Coppa Italia. On 20 June 2013, Torino redeemed the remainder of his contract from Palermo for €1.5 million.

In the 2013–14 season, he played 37 games in Serie A for Torino, who finished 7th and qualified for the Europa League after Parma failed to obtain a UEFA license. He featured in the Serie A Team of the Year, along with teammate, Ciro Immobile. On 18 May 2014, he made his 100th appearance for Torino during the 2–2 draw away to Fiorentina.

In his first appearance of the 2014–15 season, on 7 August 2014, he scored his first goal in European competition, in a 4–0 victory against Sweden’s IF Brommapojkarna, in the second leg of the 2014–15 Europa League third qualifying round. On 29 October 2014, he scored his first goal in Serie A to secure a 1–0 win over Parma, before returning to the score-sheet on 11 December 2014 in the Europa League group stage, won 5–1 away at Copenhagen. On 26 February 2015, he scored the match-winner against Athletic Bilbao, qualifying Torino for the round of 16 after a historic 3–2 victory in Spain. He concluded the season with 47 appearances and five goals between Serie A, Europe and the Coppa Italia.

Given his excellent season with Torino, on 13 May 2014, he was included in the list of 30 provisional players for the 2014 World Cup. He made his debut for the Azzurri on 31 May 2014, at the age of 24, playing as a right-back in a goalless friendly match with Ireland which was held in London. The following day, he was included in the list of 23 players selected for the 2014 World Cup. On 14 June 2014, he made his debut at the World Cup, starting Italy’s opening game against England, which Italy won 2–1. He also played the next two games against Costa Rica and Uruguay, both of which were 1–0 defeats for Italy who failed to qualify for the next round.

On 11 July 2015, Darmian signed for English club Manchester United for an undisclosed fee, believed to be £12.7 million. He signed a four-year contract with an option to extend it for a further year. He made his debut for the club against Club América during preseason on 17 July, playing the first 45 minutes, with United winning 1–0. He made his Premier League debut on 8 August 2015 as the season opened with a 1–0 victory against Tottenham Hotspur at Old Trafford, with manager Louis van Gaal selecting his performance as the best of the match. On 10 September, Darmian earned his first official accolade at the club, after he was voted Manchester United’s August 2015 player of the month. “It’s important to know this is from the fans,” Matteo told United’s official website. “I want to thank them. I am happy. I also want to thank my team-mates and everyone at the club for making me feel welcome.”

In early 2015, he was awarded with the Pallone Azzurro as the best player of the national team for 2014. On 10 October of that year, he scored his first international goal, which was the final one of a 3–1 away win over Azerbaijan which sealed Italy’s qualification to UEFA Euro 2016. On 31 May 2016, he was named in Antonio Conte’s 23-man Italy squad for Euro 2016. In the round of 16 of the tournament, at Stade de France in Paris on 27 June 2016, he came off the bench and set-up Graziano Pellè’s 91st minute volley to give the Azzurri a 2–0 win over defending champions Spain. On 2 July 2016, he missed Italy’s final penalty in a 6–5 shoot-out defeat to reigning World Cup champions Germany in the quarter-finals of the competition.

In his twenty-seventh league appearance of the 2015–16 season, on 20 April 2016, Matteo scored his first Manchester United and Premier League goal, a powerful volley from the edge of the penalty area, in a 2–0 victory against Crystal Palace. He was presented with the club’s Man of the Match award for his performance in this fixture and hailed his 55th-minute strike as the best goal of his career, thus far. His cross, in the fourth minute of play, had also previously forced an own goal from Damien Delaney. Darmian featured in the 2016 FA Cup Final against Crystal Palace on 21 May, coming on in the 66th minute for Marcos Rojo, helping United achieve a 2–1 victory.

Darmian’s first appearance of the 2016–17 season came on 15 September 2016, in a 1–0 defeat against Dutch-side Feyenoord in the UEFA Europa League group stage. He didn’t feature again until 20 October, when he played the full 90 minutes against Turkish-side Fenerbahçe in a 4–1 win. Darmian appeared in all of United’s Premier League games in November, playing at left-back against Swansea City, Arsenal and West Ham United. He continued to contribute to United’s unbeaten streak by featuring against Everton, Tottenham Hotspur, Crystal Palace and West Bromwich Albion in December. On 10 January 2017, he played in the 2–0 victory against Hull City in the EFL Cup semi-final first-leg, assisting the second goal with a cross to Marouane Fellaini. He also played in the second leg on 27 January, helping United advance 3–2 on aggregate.

Darmian’s appearances in the league became limited after a 1–1 draw against rivals Liverpool on 15 January, with him not featuring again until 19 March when he came on as a stoppage time substitute in a 3–1 win against Middlesbrough. On 9 April, he started for United, his first in the league since January, in a 3–0 victory over Sunderland at the Stadium of Light. In May 2017, Darmian played in both of the UEFA Europa League semi-final games against Spanish-side Celta Vigo, helping United go through to the final 2–1 on aggregate. On 24 May, Darmian started at left-back in the UEFA Europa League Final against Dutch-side Ajax, playing the full 90 minutes as United won 2–0.

Darmian made his first appearance of the 2017–18 season on 8 August 2017 against Real Madrid in the UEFA Super Cup, which United lost 2–1. His first Premier League appearance of the season was on 9 September, in a 2–2 draw against Stoke City. He didn’t feature again in the league until 14 October in a 0–0 draw against Liverpool at Anfield. While not featuring in the league, Darmian did play in the UEFA Champions League in a 2–0 win over Benfica on 31 October and in a 1–0 defeat against Basel on 22 November. On 20 December, he played in United’s 2–0 defeat to Bristol City in the EFL Cup fifth round. Darmian made his first league start since October on 18 April 2018, playing against Bournemouth in a 2–0 victory. Three days later, he came off the bench in the 80th minute to see out United’s 2–1 victory over Tottenham Hotspur in the FA Cup semi-final at Wembley Stadium. Darmian started in United’s last league game of the season on 13 May, overseeing a 1–0 win over Watford at Old Trafford. He was left on the bench for Manchester United’s 1–0 defeat to Chelsea in the FA Cup Final on 19 May. After falling out of favour with José Mourinho, Darmian only managed eight league appearances the whole season, five of which were starts.

Despite speculation that Darmian would depart Manchester United in the summer for more playing time, he ended up staying for the 2018–19 season. He started United’s first game of the season on 11 August at right-back in a 2–1 victory over Leicester City. Darmian’s chances were limited until 24 November, when he started against Crystal Palace in a 0–0 draw. On 5 December, he started at left-back in a 2–2 draw against Arsenal. He played at right-back in United’s 3–1 defeat to Liverpool on 16 December. On 5 January 2019, Darmian played at centre-back in Manchester United’s 2–0 FA Cup third round win over Reading. Caretaker manager Ole Gunnar Solskjær praised Darmian’s performance by saying, “He was one of the few who settled us down and tried to pass it and I don’t think he put a foot wrong.”

On 2 September 2019, Darmian returned to Italy signing for Parma on a four-year deal.

On 5 October 2020, Darmian joined Inter Milan on a season-long loan; Inter have an obligation to make the transfer permanent for £1.8 million at the end of the loan period.