Virgin Vision

One character has recently lost their virginity (or a pair of characters has recently lost their virginity together). This fact is immediately obvious to yet another character (usually, but certainly not always, an older woman), despite there being absolutely no outward sign or obvious reason to think that. The characters aren't acting cute together, there's no tell-tale messy hair or mussed clothing, nor even the subtle aroma of "just had sex".

The other character merely looks upon the new non-virgin, as if using Virgin Vision as a superpower, and they know. (Women's Mysteries, I suppose.)

Compare to Did You Just Have Sex? (where other people know because the characters are giving away clues) and Virginity Flag (where there is some obvious means of telling who is and is not a virgin).

Examples of Virgin Vision include:

Comics

As an inversion, in the French comic Le Chant d'Excalibur, everybody can apparently detect that the heroine is still a pucelle, and will mention it at the first occasion.

Film

In Shakespeare in Love, Queen Elizabeth can tell that Viola has been "plucked since [she] saw her last." The Queen adds that "it takes a woman to know it," though she doesn't reveal how, exactly.
This happens to the main character in Real Women Have Curves. Her mom figures it while watching Ana stand in front of a full-length mirror.
In the West Side Story movie, Anita doesn't say it out loud, but she gives Maria this look that says it all.
The second Ace Ventura film:

"She's not a virgin!"
"They can tell that?"
(Cue being Chased by Angry Natives)

In Say Anything, Lloyd Dobbler's friends can tell that he and Diane Court "did the deed", despite Lloyd's "I admit nothing" disclaimers.

Literature

In the Morganville Vampires, Claire fears this trope. Sure enough, Eve barely looks up and realizes what happened.
In The Sound of Waves by Yukio Mishima, older women claim to be able to determine whether a girl is a virgin just by looking at her breasts.
In The Dresden Files series, White Court vampires can sense virginity, which led to a Crowning Moment of Funny when Lara Raith (leader of the White Court vamps) met Carlos Ramirez (a Cassanova Wannabe).
- Justified; they feed on energy, most particularly sexual energy.
The first chapter of Alina Adams figure skating mystery Death Drop starts with the researcher heroine listening to commentator insisting "You can always tell a virgin by the way she skates." She thinks the non-virgins skate better. She turns out to be right in her assessment of the skater they were watching.
In Blue Smoke by Nora Roberts, Reena's mother is able to detect instantly that her daughter has lost her virginity when Reena arrives home after doing the deed. The scene is retained in the made-for-TV movie based on the book.

Live Action TV

After Donna and Eric finally have sex in the third season of That '70s Show, neither one admits anything to the other characters (despite Eric really wanting to do so). When Donna goes to talk to Jackie, Jackie see her as having the words "I Had Sex" written on her forehead, and Jackie immediately rushes her away so they can talk about it. When Eric goes to talk to Bob, Bob sees Eric as having the word "Guilty" on his forehead, inidcating that Bob is suspicious, but Bob makes no sign of knowing what happened until he officially finds out a few episodes later.
In the Buffy the Vampire Slayer episode "Innocence," Buffy's mom can tell there's something different about her daughter, but not what. Buffy was acting as if she had something to hide, but that's hardly unusual for her.

Video Games

Oghren in Dragon Age claims he can "smell purity". Apparently it's not a very useful skill.

Web Comics

This strip from Sabrina Online.
Combined with Aura Vision, it is a superpower in the Ciem Webcomic Series. Leaves the heroine feeling Defiled Forever, though she does adjust to it eventually.