Assassin's Creed: Syndicate is the ninth major game in the Assassin's Creed series and like previous titles it is a third-person action game with stealth set in a large open world environment. This game takes place in London in 1868 during the Industrial Revolution. Like many of the earlier games it is told as a simulation played in the present using the Animus technology from the Abstergo company from the viewpoint of an anonymous Assassin initiate. In the present Rebecca Crane and Shaun Hastings clash with the Templars who are searching for a Piece of Eden in modern day London. There are regularly short sequences in modern times, but the game is almost entirely played in the simulation of London following the Assassin twins Jacob and Evie Frye who look for Pieces of Eden. Their main target is also to thwart the Templar Grand Master Crawford Starrick who rules all of London by day and uses the Blighters street gang at night to get his dirty work done. Jacob and Evie need to bring him down by influencing and infiltrating the criminal underground, but also need to take over London with their Rooks gang fighting the Blighters, one area at a time.

After the initial sequences players can freely swap between Jacob and Evie at any moment in the open world, a first for the series, but they each have their separate main missions. Evie favours a stealth approach and is mostly after the Pieces of Eden, while Jacob is more confrontational in his actions and mainly wants to bring down Starrick. In the background many other Assassins are at work to stop the Templar control over the city. Most side missions are open to any character, but the main ones require a specific character to complete them. The characters have their separate storyline, but these often intersect. There are some separate missions that take place in a very different time as well, due to an Animus glitch that opens rifts in time. Many famous historical characters are a part of the story, such as Karl Marx, Charles Darwin, Florence Nightingale and Charles Dickens. There are also iconic buildings and environment such as Westminster Palace, the Big Ben, the Bank of England, the Tower of London and Buckingham Palace. The characters share money, experience and rewards, but have separate gear and skills. Like earlier titles a currency and experience is earned to buy and upgrade gear and weapons. Experience is converted into skill points that can be invested in three skill trees: combat, stealth and ecosystem.

As with previous game exploration and climbing is encouraged through a free running system. It is similar to the one in Assassin's Creed: Unity with two options to pursue a path up or down. Entirely new to the series is the use of moving trains and driving through London using horse-drawn carriages, including one as a mobile headquarters. This is used to transport people or goods, or chase enemies, but it is also possible to kidnap enemies, put them in a carriage and demand a ransom. A rope launcher can be used to rappel up buildings quickly and it can also used as a zipline (returning from Assassin's Creed: Revelations) to get down. There is a separate stealth system based on a ring and eagle eye vision. Fighting is done using close combat melee attacks with fists, brass knuckles and canes. There are also ranged weapons with many types of pistols. The system to punch, attack and block/counter is fairly similar to previous games. Enemies can now be beaten into a 'near death' status where they remain defenseless. When this is done for several enemies, there is an opportunity for spectacular multi-kill finishing moves and executions. With a later skill, performing these can also scare other enemies away.

Next to the main missions there are various sidequests and activities to earn additional money such as carriage races and fight clubs. Many additional missions are based around territory control, taking over London sector by sector, similar to the gameplay of Assassin's Creed: Brotherhood. This is done by completing activities and killing rival gang members. To completely secure a sector a large-scale gang war needs to be fought. The presence of Rooks in the area then becomes immediately visible and it is possible to invest in various gang upgrades to make them stronger. Unlike the previous titles back to Brotherhood, there is no multiplayer component to the game. Compared to the previous game the leveling system has been simplified, there are less currencies and there is no (required) integration of a companion app to unlock certain content. A few extras can be unlocked by logging into Ubisoft Club. Optionally the in-game currency can be bought with real money.