Holmes and Watson arrive at the Roylott's house, where Helen is waiting for them. Holmes warns her that her stepfather knows that she consulted him, and that she must be on her guard. Holmes begins his investigation, examining the house. He notes that while a lot of the building is in bad condition, the wing which Dr. Roylott and Helen live in is in pretty good shape. He mentions this to her, saying that it doesn't seem like this section of the house needed repairs. Helen tells him that it didn't; she suspects that it was done to force her to move into Julia's room. Holmes tests the window on this room, and finds that it's impossible to break in from outside when the metal bar is locked on the inside. This ruins his theory of the doctor having a gypsy accomplice gain entrance in that way. With that possibility eliminated, the investigation moves indoors.
Julia's room is pretty bare, but it does have a couple of features which interest Holmes. One of these is a decorative bell pull which Helen says was installed only a couple of years ago. When Holmes tries it, he finds to his surprise that it is a fake, just attached to a hook rather than a bell wire. Close to the bell pull on the wall is a ventilator which runs from Julia's bedroom to Dr. Roylott's. Holmes finds this strange, as it would make more sense for a vent to run to the outside for fresh air, rather than another room. Helen tells him that the vent was installed around the same time as the bell pull. Holmes asks her to let them have a look at Dr Roylott's room.
Roylott's room is also sparsely furnished, but does contain a safe which Holmes finds interesting. He asks Helen what it contains, and she says that she thinks it has her stepfather's private papers in it. Holmes asks her if there could be a cat in it, pointing out that there is a saucer of milk sitting on top of the safe. But Helen says that the only cat on the premises is the cheetah. Holmes' interest is then drawn to the chair in the room, and he carefully examines the seat cushion on it. The final item which catches Holmes' eye is a dog leash with a small loop tied at the end. Watson and Helen are at a loss to make any sense of all this, but Holmes appears to have settled in his mind what is going on. He grimly tells Helen that she must follow his advice to the letter, because her life may depend on it. She promises to do whatever he says. Holmes tells her that he and Watson are going to take a room at the nearby inn, from which they can see the manor. Meanwhile, when Dr. Roylott returns home, Helen is to plead a headache and lock herself in Julia's room. When she hears her stepfather retire for the night, she is to unlock the window and put a lamp in it as a signal. She is then to slip quietly to her old room and spend the night there. When they see the light, Holmes and Watson will sneak over to the house, climb in the window, and spend the night in Julia's bedroom. Helen begs Holmes to tell her what he suspects happened to her sister, but he tells her that he wants to obtain some proof first. Then, after encouraging her to keep her courage up, the two men leave for the inn.
At the inn, they engage a room which overlooks Roylott's house. As they watch, they see Dr. Roylott arrive home late in the evening, obviously still in a rage. Holmes tells Watson that he has some reservations about taking him along with him, since there's a distinct possibility of danger. But Watson tells him that as long as he can be of help, he's coming. He then questions Holmes about what he saw in the two bedrooms that no one else noticed. Holmes tells him that he knew before they left London that there would be some sort of vent between Julia's bedroom and Roylott's from Helen's description of Julia smelling their stepfather's cigar smoke. He asks Watson if he didn't find the fake bell pull and the oddly placed ventilator significant, especially since they were put in at about the same time. He also tells his friend something else he noticed: Julia's bed was bolted to the floor, meaning that she couldn't move it and it would always be in close proximity to the vent and rope. Watson begins to get an inkling of what Holmes is suggesting, and is horrified. Holmes says that when a doctor goes bad, he makes the cleverest criminal because he has knowledge and nerve. Around eleven pm, the lamp appears in the bedroom window and Holmes and Watson sneak to the house, crossing the lawn without detection, though they have a bad moment when they run into the baboon.
Once inside, they turn out the lamp and sit in the dark. Holmes whispers to Watson that he must not fall asleep, because his life may depend on his being alert. He has armed himself with a long cane, and tells Watson to keep his gun handy. The hours pass as they sit silently in the darkness until, around three am, there is a very brief gleam of light from the vent, and the smell of lamp oil. Watson realizes that someone has lit a dark lantern- a lamp with a metal panel to mask the light. Another half hour passes, and Watson hears a slight hissing sound, almost like a kettle. Holmes leaps into action, lighting a match and then using the cane to strike furiously at the bell pull. Watson, blinded by the sudden light after hours of complete darkness, can't see what Holmes is attacking, but does hear a low whistle such as Helen had described. As his eyes adjust, he can see the look of "horror and loathing" on Holmes' face. Holmes stops hitting at the bell pull and stares up at the vent. Suddenly, they hear a horrible shriek coming from Roylott's room, which strikes horror into the hearts of all who hear it. Watson, gasping, asks Holmes what it means. Holmes says that it means that it's over, and cautiously enters the other bedroom, telling Watson to keep his pistol at the ready.
In the doctor's bedroom, they find a hideous sight. In the light from Roylott's dark lamp, they can see the door to his safe is wide open. Roylott is sitting in his dressing gown in a chair, his dead eyes open and fixed on the ceiling. There is a strange yellowish band with brown spots wrapped around his brow... the speckled band- a.k.a. the swamp adder, the deadliest snake in India. As they step closer, its head emerges from the doctor's hair, and Holmes grabs the dog leash with the loop in it, lassos the snake, and throws it back into the safe.
The morning is extremely busy as they pack the shocked Helen off to stay with her aunt and deal with the authorities. It is not until they're on their way home that Holmes has the opportunity to explain everything to Watson. He says that the vent and bell pull combined with the immovable bed made him suspicious that something was being sent through the vent to climb into Julia's room. Knowing that Roylott had brought several exotic animals from India, he hypothesized that he had a poisonous snake. A medical man, Roylott picked one with a fast acting and virtually undetectable poison and trained it with milk to come to his whistle. He would put the snake through the vent, it would crawl down the pull, and land on the bed. Holmes explains that the chair cushion in the doctor's room had footprints on it, where he had stood to reach the ventilator. The adder might not bite the bed's occupant the first or second time Roylott let it in, but eventually the bite would occur, and the victim would die. On the night of Julia's death, Helen heard Roylott whistling for the adder, and then the metal clank of the safe door as he shut the snake back in. When Holmes was striking the snake with his cane, it escaped back through the vent, enraged, and attacked the person on the other side- Roylott. Holmes says that, in a way, he is responsible for Dr Roylott's death, but that it's unlikely to weigh too heavily on his conscience.