The Wait Until Dark scene was the first that came to my mind.
Likewise, the cold open to The Spy Who Loved Me. The Union Jack was icing on the cake.
Saw The Sixth Sense in a packed theater about a week after it opened, so before spoilers were widespread. Enormous collective gasp at the reveal of the big twist.
Everyone talked about sitting in stunned silence at the end of Avengers: Infinity War. Guess folks weren’t expecting the bad guy to win.
In the film Sorry to Bother You, which has many WTF moments, there’s a scene where the black MC is at a party with a bunch of white people and they pressure him into performing a rap.
Well, he raps. He screams “N**** shit! N**** shit! N**** n**** n**** shit!”
And the white people enthusiastically shout it right back.
Apparently the white extras were so horrified by this scene they didn’t want to do it. It definitely ranks as among my top most uncomfortable moments in film.
We were at the movie Shazam! and nearing the climax when a little boy behind us piped up, "Oh, no, he’s going to lose just like the Avengers!
That movie messed kids up.
I showed my brother a movie he hadn’t seen before but I had: One Flew Over the Cuckoo’s Nest.
When the Chief said, “Thank you”, my brother just about fell off the couch. 
The ending of “The Mist” caught me off guard completely. That was not expected after a fairly mundane and cookie-cutter horror movie. Especially since the Stephen King short story ended so much differently.
I had heard the spoiler but didn’t see the movie for so long that I forgot it and it worked quite well on me.
That was one of Tarantino’s most brilliant moments. He knows what audiences have been trained to expect, the moment of silence, fluttering of eyelids, gradual awakening… and turned it into a shot of adrenaline directly into our own hearts.
Surprised no one has mentioned it yet, but the ending of The Usual Suspects.
“And like that…he’s gone.”
“Seven”- What was in the box.
I saw Pulp Fiction at a Screen Actors Guild screening, so this was a very sophisticated audience used to seeing absolutely everything. There were people literally jumping up out of their seats and shouting.
Planet of the Apes…
Yeah, I was just a 12 yr old kid, but damn that ending hit me hard.
After enjoying an hour of a nice but kind of silly movie, suddenly in the last minute there’s a whole new meaning to think seriously about. .
And the way it comes on slowly…the gradual reveal (starting with just a shadow, then the full statue), is good cinematography…
'Twas an excellent scene !
And for any who don’t know the back story … it’s equally cool !
Same movie - different scene. When Bruce first surfaces and smiles at Chief Brody, I dropped my beer! (Saw it at a drive-in.)
I’ll add The Line from Shark Attack 3: Megalodon. Completely unexpected, but anyone who watched the DVD immediately rewound to see if they really heard what they thought they did.
Not long after the release of Seven (SE7EN) I was at a friend’s housewarming party.
When they were opening gifts, he shook a box, opened a bit of the top, looked in, and I SCREAMED, “WHAT’S IN THE BOX???”.
Three people had seen Seven, and cracked up. The remaining 20 or so guests didn’t have a clue.
Just that line got an immediate (laughing) response from me. I didn’t even have to click on the link!