The class is studying nocturnal animals at school and eventually checks out the attic of the classroom. Inside the attic, Tim notices a bat hanging from the ceiling and confirm that the bat is the only nocturnal creature they know very little about. Just then, the bat flies away and Ms. Frizzle suddenly appears, dressed as a bat. She tells the class that she will "show them the secrets of being nocturnal". As Ms. Frizzle tells the class more about the night, Ralphie becomes apprehensive.
The students' parents arrive at the school, and Ms. Frizzle, after being introduced to Keesha's grandmother, tells her that she could just "eat you up", exacerbating Ralphie's trepidation. When Ralphie's mother Dr. Tinnelli meets Ms. Frizzle, the latter states that, "I do so enjoy taking him under my wing", which finally convinces Ralphie that Ms. Frizzle is a vampire as he tells the rest of the class. He uses Ms. Frizzle's term "The darker the night, the safer the flight" as evidence to support this, but Dorothy Ann states that Ms. Frizzle was talking about bats, for when it's dark, they blend in and are unobtrusive to enemies. Ralphie, however, is not convinced as he states that vampires come out in the night to. The conversation is interrupted when the class overhears Ms. Frizzle informing the parents that the class and their parents will go someplace more "suitable" and "breathtaking" right before letting out a sinister laugh. As the parents enter The Magic School Bus, Ms. Frizzle tells them that the children "will be taking care of" and takes them on a nighttime field trip to learn more about bats. The class follow her and their parents via 8-seat bat mobile driven by Liz to a scary old castle.
As they arrive at the castle, they spy on Ms. Frizzle and their parents, who enter the castle. When Ralphie keeps trying to convince the class that Ms. Frizzle is a vampire, Keesha, having enough of this, decides to see for herself and the class walks to the castle. Wanda tries to open the door. But the handles break apart. When Ralphie views Ms. Frizzle and the class' parents drinking a beverage each through the window, he becomes convinced that Ms. Frizzle is making the adults drink blood.
Keesha tells Ralphie that the adults are drinking tomato juice, which Ms. Frizzle served at the class picnic. Ralphie then comes with a plan to drive a textbook in the heart of who he calls the "teacher vampire". When Carlos swats mosquitoes off of him, the bats fly around the class, and Phoebe tells the class that the bats are really eating the mosquitoes, causing Keesha to come to the conclusion that the bats aren't vampires.
Meanwhile, Ms. Frizzle gives the adults a tour through the inside of the castle and they walk around "bat guano", the waste of the bats that makes great fertilizer according to Ms. Frizzle. When they walk through a stuffy hallway, Ms. Frizzle tells the adults that it's from the breath of the bats, and Mr. Ramon jokingly states that "bat breath is better than no breath at all". They then come across pink bat progeny, which Ms. Frizzle calls "the children of the night".
The class also views them through the window and Keesha informs Ralphie that the bat babies drink milk and not blood. Ms. Frizzle tells Keesha's grandmother (who has heard Keesha's voice from outside) about how bats protect their young and Ralphie is convinced that Ms. Frizzle will feed the class to the bats, misconstruing tidbits (food fed to bats) as "kid bits". So he brings the class with him to get help.
When they enter The Magic School Bus, Ralphie tries to drive it so the class can get out of here before Ms. Frizzle catches them. He pushes the bat button and The Magic School Bus transforms into a bat, which flies through the woods. The class are impressed of the fact that The Magic School Bat isn't running into any trees. They notices that as The Magic School Bat makes the pinging sound, it opens its mouth and wiggle its ear to listen to the echo of each ping. Keesha finally infers that when the sound the bats make echo back, they know something is there and avoids crashing into it, as they use their echoes to locate things. They also learn that bats see more with their ears rather than just their eyes. When The Magic School Bat eats moths, the class also learn that bats are at night to find food in the dark. Ralphie presses another button on the bus, which transforms him and the rest of the class into bats. Just then, they come across an owl, who attempts to catch them, only to miss in the process. The students then learn that the bats' enemies can't see them due to the dark.
When the students overhear a vociferous Ms. Frizzle stating, "At last, the moment I've been waiting for has come", Ralphie tells the class that the vampire in his comic book said the same thing and the class fly to the castle, with Keesha reluctantly starting to believe Ralphie's theory. They hear Ms. Frizzle asking who would be her first victim, and Keesha, after overhearing her grandmother telling Ms. Frizzle to take her, flies inside the castle, shouting, "DON'T YOU DARE BITE MY GRANDMOTHER!"
The rest of the class and The Magic School Bat follow along and use their sound to locate Ms. Frizzle and the other adults. They come across more bats and finally hear the adults voicing their opinion of the field trip to Ms. Frizzle, as Keesha's grandmother tells the teacher, "I can't believe I let you do this to me" and Dr. Tinnelli states that, "My neck will be sore for a week". Ralphie brings the class with him to fly to Ms. Frizzle's location as Mr. Perlstein is heard saying, "Maybe we should've stayed home tonight".
The class finally spot Ms. Frizzle and the rest of the adults hanging from a chandelier dressed in bat costumes. The class fly through them and the adults extricate themselves off the chandelier. When the class finally find the truth, they all chide Ralphie for his theory. Ralphie apologizes to them and acknowledges the fact that bats aren't vampires, but nocturnal creatures that come out at night to feed on insects. Carlos jokingly states that "The bat news is we're still bats", to which Phoebe jokes "Fangs for reminding us, Carlos!", causing everyone to groan and laugh "Phoebe!". Liz comes around with The Magic School Bat and the class hops inside to transform to their human form and The Magic School Bus back to normal, just in time for their parents to exit the castle. When Dr. Tinnelli asks Ralphie what was going on, Ms. Frizzle states that he and the rest of the class were just "hanging around", with Ralphie stating "bats all" as he and Ms. Frizzle laugh.
After the story, Ms. Frizzle names the bat Belinda so she puts in the bag. But Katie explains that if she only puts scary things in the bag, then she won't put the bat in there. In fact, it's kinda cute. Ms. Frizzle agrees and as her aunt Bodacious always says "The more you know something, the less you fear" and puts the bat back up in the ceiling where she belongs. Richie asks Howard if that guys reminds him of somebody and Katie ask the Friz what else she got in the bag. Ms. Frizzle says nothing and as she begins to leave, Richie, Howard, and Katie asks anything such as what she did but she says that she didn't do anything and the students actually did. Ms. Frizzle says that the students began this victim of their own fears but they took chances, made mistakes, and they conquered them. Just then, the Magic School Bus beeps his horn and Ms. Frizzle leaves the museum and calls out "Bus, do your stuff!" The students open the door and see Liz inside the Magic School Bus and Ms. Frizzle drives the bus off while saying "Wa-ha-ha-ha-hoo!" After the bus leaves to Walkerville, the students all say "Wow, we've been frizzled!" and they give hi-5 while jumping through the air in conclusion.