Hidamari Sketch. A girl's father finds a short hair not of his daughter's color while visiting her home and wonders if she has a cat. Before anyone answers, a Bifauxnen walks in and he immediately accuses her.
One single-page Archie comic has Archie inexplicably the First Teen on Mars. He's asked a bunch of questions regarding the terrain, the weather, etc. Cue Ronnie with, "I have one question. Whose purple hair is this?"
Parodied and played straight in The Incredibles: Helen plucks a piece of rubble from Bob's jacket in order to confront him on his late night vigilantism. Later she discovers Mirage's platinum blonde hair on his superhero uniform and begins to grow suspicious.
A scene that was recorded but not animated has her confronting him about this. He claims that he had the uniform taken to the dry cleaners (passing it off as a costume for a party), and the hair comes from the old lady there.
Murder by Death: Sidney Wang tries to one up the detective group's Sherlock Scan by concluding that Mrs. Charleston must have dyed her hair red, because the hairs on Mr. Charleston's jacket are blonde, only to realize the truth halfway through his sentence.
In That Old Feeling, Rowena's bright red hair is a significant part of the backstory because it proved that she'd had an affair with Molly's father while he was still married to Lilly, Molly's mom. At the climax of the movie, Rowena's bright red hair also reveals that she'd slept with Molly's newlywed husband on the previous night.
In the film Crossing Delancey, an African-American character with curly black hair finds a long, straight blonde hair on her boyfriend, which she takes to mean that he's cheating on her.
Usually affair hair is found in the bedroom, and belongs to the "other woman," but The Kids Are All Right uses a variation, when Nic discovers the truth after finding her wife's distinctive red hair in a bathroom drain at a friend's house. The woman's excuse of having taken a shower to clean up after working in his garden falls flat when Nic reveals that she found her hair in his bed as well.
So this woman says to her friend, "I'm worried my husband might be cheating on me, but I don't know what to do because there's no evidence." "Oh, that's easy," says her friend, "You just check the lapels of his jacket when he gets home, and then if you find a blond hair you say 'Oh, so you've been seeing a blond woman!' Or you find a brown hair and say, 'Oh, so you've been seeing a brunette!' Then he'll have to confess." The woman thinks this is a great idea, so when her husband comes home that night, she checks his jacket. There's no hair on it at all. So she charges in and yells, "Oh, so you've been seeing a bald woman!"
A Rose for Emily ends with a strand of gray hair found next to a body that's been dead for decades.
In Wizard and Glass, Roland's friends discover that he's sleeping with Susan this way.
In the Indian novel The White Tiger, this is how Balram realizes that Ashok slept with the Ukrainian student.
Live Action TV
In Young Dracula, they have some fake Affair Hair - Vlad gets some of his stuffed wolf's hair and pretends it's from his mother's werewolf boyfriend.
This happens in the pantomime sketch "Argument to Beethoven's 5th" by Sid Caesar and Nanette Fabray, just as the couple seems to have made up. At the very end, she figures out that it belongs to the cat, and they're reconciled.
In My Name Is Earl, Earl finds a piece of Darnell's hair on his pillow, but believes it's his own hair. He does find out about the affair nine months later, though.
Played with in Roseanne. Roseanne notices a hair on one of Dan's shirts, and is suspicious for a moment, before realising it's actually one of hers.
Parodied in Paul Merton's TV Show where Paul's wife accuses him of having an affair with Gloria Hunniford on the grounds that her entire wig was in the back seat of the car. Paul reasonably points out that it was, in fact, some hay for the rabbit.
Mariah Carey's "Heartbreaker" has Jay-Z rapping about how "she found a strand of hair, longer than hers". The video shows a toon Mariah finding an Affair Hair.
An early story arc in Foxtrot features Andy thinking Roger is having an affair, when he's really just working overtime. Andy admits she was wrong and they hug, only for Andy to pluck a hair off his jacket and ask "Whose hair is this?"
For those who are not familiar with the strip, Roger is balding, and in denial about it.
There was a Far Side strip that caused a bit of controversy because it showed a female chimp picking a blond hair off her chimp husband and asking if he'd been doing more "research" with "that Jane Goodall tramp". Jane Goodall, by the way, loved that strip.
Jane Goodall herself told the story on the title page of one of the Far Side collections. Her society was getting ready to sue over the strip when she returned from field work. Her reply was, "Wow, real fame at last!" Now, it's one of her society's biggest moneymakers, appearing on many fund-raising products. Ms. Goodall invited Mr. Larson to her preserve - where he promptly got the snot kicked out of him by her lead chimp, a male named Frodo. Just as Planned?
Catherine has a variant, as it's the girl Vincent's cheating with that finds his girlfriend's hair. Vincent also explains the long pale hair as coming from his chest. In another twist, she knew Vincent had a girlfriend the whole time.
Sabrina Online has a gag along these lines after Sabrina's dad has the tour of Zig Zag's porn studio - while the hair color is nothing to worry about (since both he and Zig Zag are skunks), the length is...