While the film is far more serious than the Dollars Trilogy, constant sarcasm from Harmonica and Frank occasionally crosses the line into hilarious.
Frank: How can you trust a man who wears both a belt and suspenders? The man can't even trust his own pants.
Guess where Frank shoots the informer when the idiot proves to be too much of a liability? Hint: Frank shoots him three times...
And Cheyenne's reaction to the informer's death is deadpan: "Told you to keep your mouth shut."
Also an in-joke. Leone himself wore a belt and suspenders.
Some scenes in the 11 minute intro, probably an intentional example. And the tense and cool exchange afterwards.
Harmonica: Did you bring a horse for me? Frank's Man: Looks like we're shy one horse. Harmonica: (shakes head) You brought two too many.
Harmonica: I saw three of these dusters a short time ago, they were waiting for a train. Inside the dusters, there were three men. Cheyenne: So? Harmonica: Inside the men, there were three bullets.
This little gem:
Harmonica: The reward for this man is 5000 dollars, is that right? Cheyenne: Judas was satisfied with 4,970 dollars less. Harmonica: There were no dollars in them days. Cheyenne: But sons-of-bitches? Yeah.
Morton: Tell me, was it necessary that you kill all of them? I only told you to scare them. Frank: People scare better when they're dying.
Cheyenne doesn't care one iota about the final duel and is just annoyed when the shooting makes him cut himself while shaving.
The exchange between Jill and a barman.
Barman: I can tell you're accustomed to fine living. Bet you come from one of those big Eastern cities.
Jill: New Orleans.
Barman: New Orleans!
Jill: You've been there?
Frank's henchman Snaky's reactions to a fly landing on his face that he doesn't even bother to swat away with his hand. Finally he traps it in a gun barrel. When it stops buzzing inside, he checks to see if it's dead, only for it to start buzzing again.
In the same scene, a residue from the ceiling keeps dripping onto Stony's head, repeatedly. He puts on his hat and the water keeps dripping onto his hat instead.
The Native American woman who is chased out of the station mutters to herself as she runs, "Dumb palefaces..."