The game begins as Eric Cartman runs over to Stan's house to warn him that 'horrible beings' are attacking South Park. The two are able to defend the household from an army of Ginger Kids and Cows before picking up Kyle Broflovski and running to their school. Once again, they manage to protect the area, but Cartman receives a phone call from Kenny McCormick, who is under attack. After helping Kenny to fight off another horde of enemies, the boys deduce that the one responsible for the mayhem can only be an evil supervillain: Professor Chaos.
The boys reach Butters Stotch's house and confront Chaos, but he claims that his only evil plan was 'to replace all the healthy mineral water in the pharmacy with boring old regular water'. During a battle with another group of enemies, one of Butters' testicles is shattered, so the boys take him to Hells Pass Hospital and protect it while he is healed. Jimmy Valmer runs in to tell the boys to head to Stark's Pond, where they help Jimbo Kern and Ned Gerblansky to stop another group of enemies from destroying the town. Craig Tucker directs them to the lair of the The 6th Graders, led by Scott Tenorman, who have been recruited by the sinister evil that looms over the town.
After defeating Scott and the 6th Graders, Clyde Donovan tells the gang to help Mr. Lu Kim to defend the Great Wall of South Park from an army of Mongolians. After a long battle, the boys and Lu Kim are victorious, but only before being redirected once again, this time to the South Park Docks, by Tweek Tweak.
The boys defend the Docks from a group of enemies led by General Disarray. After doing so, Pip Pirrip attempts to reveal the villain's identity to the boys, but, typically, they reject him before he is able to finish his sentence. The boys head up high into the mountains above South Park to meet Randy Marsh, who tells them that they are about to be attacked. They are able to stop ManBearPig and a mass of Demons from taking over the geology building, before Timmy comes to meet them and they are directed one final time to Downtown South Park. Randy confirms that they must head Downtown to stop the villain, and warns the boys that 'he has always been waiting' for them.
The boys stop a huge army of assorted enemies from destroying the town hall before a voice announces 'Game Over' and the game world collapses. After the boys are pulled out into a world beyond the Universe, the villain is revealed to be the Japanese Announcer (from the episode Chinpokomon), who has been manipulating South Park and using the boys as pawns. The Announcer then attacks South Park and attempts to storm the town hall himself, but the boys are able to defeat him. Without an ultimate villain controlling South Park, the evil forces disappear and everything returns to normal.
Like most tower defense games the goal is to try to stop enemies from crossing the map by building towers which shoot at them as they pass, however this game is unique as, instead of just towers, the player can also steer him/herself around and attack with snowballs, which is a change to most tower defense games. Enemies come 3 to 16 waves of foes that you have to defeat and there are bosses to fight at the end of each of the five last levels. The enemies are usually characters that the protagonists have fought in the show before, or major 'villains' from the show, with the exception of the final boss.
- Eric Cartman - His special ability is causing explosions to occur around the field similar to Professor Chaos
- Kenny McCormick - Causes enemies to drop up to 20 coins without needing to be defeated
- Stan Marsh - Partially heals towers and heals the town for 20 points
- Kyle Broflovski - Increases everyone's speed and damage for the next 7 seconds. Many gamers use Kyle for defeating bosses
- Butters Stotch - Spins around incredibly fast, forming a tornado around him that kills anything it comes into contact with, as the song "I've Got Something In My Front Pocket For You" plays.
- Jimmy Valmer - Similar to Butters, only he twirls his crutches around
- Craig Tucker - Slows down enemies for 20 seconds.
- Clyde Donovan - Throws a colostomy bag which explodes on nearby enemies
- Tweek Tweak - Similar to Butters only to flaming killing everything he sees and last only for 5 seconds
- Pip Pirrip - In a rage, Pip can pummel through enemies at a slightly increased speed
- Timmy Burch - Similar to Butters but is faster and is electrical based
- Wendy Testaburger - Halves the price of towers for 10 seconds. Unlocked upon completion of the game.
- Bebe Stevens - Automatically picks up all coins on screen. Unlocked upon completion of the game.
- Red - Summons a rainbow while stars rain down inflicting medium damage. Unlocked upon completion of the game.
- Token Black - Plays jazz music, "depressing" enemies, inflicting medium damage. Unlocked upon completion of the game.
- Professor Chaos - Unlocked through a special unlock code. One may receive the code if the developers think you're good at the game. His ability is causing Chaos to reign all over the field.
- Gnomes - Gnomes steal underpants in an attempt to profit off them somehow. Gnomes are vulnerable to Lasers and do 5 points of damage to the town if they get through. They are also very fast.
- Ginger Kids - Cartman's prejudice turned these normal-looking kids into deadly creatures. Ginger Kids are largely considered to be the weakest enemies in the game. They are slow, fragile, only do 1-3 points of damage and are effected badly by all towers. However, on insane difficulty, they'll appear in the hundreds, nearing the two hundred, making them more threatening.
- 6th Graders - A group of older kids who pleasure themselves on beating up 4th graders. 6th graders are the only enemies - besides Mongolians to have 2 types. This type does 5 points of damage, move at an average pace and take average damage from all towers.
- 6th graders on bikes - A faster version of the 6th graders, they use their bikes to speed right past your defenses. They are fast, do 9-11 damage to the town and are most effected by fiery-works towers.
- Christmas Critters - Cute little critters that revealed themselves to be devil worshipers after tricking Stan into helping them. They are fast, do 9-11 points of damage and take most damage from fiery-works towers.
- Jakovasaurs - Near-extinct creatures that annoyed everyone in South park. - Except for Eric Cartman, who found them quite amusing. Jakovasaurs do 9-11 points of damage, move slowly and are most vulnerable to Pee-Tron Towers.
- Mongolians - Tuong Lu Kim's sworn enemy, they constantly broke down his city wall and abducted the children of South Park. Mongolians come in two types. These ones are slow, do 9-11 points of damage and are most vulnerable to cherry bomb towers.
- Terrorists - British soldiers who waged war on South Park. They are slow and take extra damage from cherry-bomb towers, but are very durable and do 9-11 points of damage to South Park.
- Homeless People - These folks came to South Park after Kyle gave one of them 20 bucks he was saving up for a Xbox game. They move at a moderate pace, take normal damage from all towers and do 5 points of damage.
- Demons - Direct from hell, Demons are the minions of Lord Satan. They are slow and take great damage from pee-tron towers, but do 9-11 points of damage to South Park if they get through.
- Crab people - Evil creatures that tried to metrosexualise man to weaken them and take over the planet. They are fast and do 9-11 points of damage if they get through, but are weak against fiery-works towers.
- Zombies - Undead beings who sustain themselves on the flesh and brains of the living. They are slow, do 9-11 points of damage if they get through and are weak to the pee-tron tower.
- Hippies - Cartman's secret phobia, they have a tendency to create drum circle in a person's backyard. They are average in speed and toughness, doing 5 points of damage and taking normal damage from all towers.
- Yanagopa - or pygmies - They miniscule creatures hail from the Amazon rainforest, fighting off wild animal with their spears. They are fast, but only do 5 points of damage to South Park and are easily eliminated by Lasers.
- Cows - The Humble livestock of South Park. They are slow, drop 3 coins, as opposed to the singular coin that other enemies drop, deal 1-3 points of damage and only appear on the first level: Stan's House. They appear in small numbers.
- Old People - The Elderly citizens of South Park who once tried to take over the town. They are amazingly tough and do 9-11 damage but are slow and weak to cherry bomb towers.
- Fast pitch - Throws baseballs at a high speed to hit enemies. (Equally effective on all enemies)
- Laser - A weak, but fast, laser that can hit multiple enemies. (Most effective on Gnomes and Yanagopa)
- Cherry bomb tower - A tower that launches a cherry bomb that splits into smaller parts. Enemies who are hit are set on fire and take damage over time. (Most effective on Old people, Mongolians and terrorists.)
- Fiery-works tower - This tower fires rockets that lock on to enemies and home in on them. (Most effective on Crab people, Critters and 6th graders on bikes.)
- Pee-Tron tower - This tower uses a cat that heavily urinates to shoot poison shots that slowly drain their targets of health. (Best used on Jakovasaurs, demons and zombies.)
- Refrigerator tower - A tower designed to freeze enemies in their tracks, thus leaving them open to attack from other towers, stage hazards, or players. (Works on anything except bosses.)
- Sticky bomb tower - This tower launches a bomb that latches itself onto to a single enemy and explodes after 3 seconds. (Effective on all enemies, must be unlocked by completing challenge level 4.)
- Plasma ball tower - A tower that shocks everyone near it with electricity, rendering them vulnerable. (Effective on all enemies, must be unlocked by completing challenge level 5.)
Bosses are extra tough and are unaffected by Refrigerator towers. If they enter the town then it's game over. The exception is Mongolians on Horses who make up for it in speed and numbers.
This is a list of achievements that can be earned in South Park Let's Go Tower Defense Play!.