Prepare your tear ducts.
"We can't fight nature, John. We can't fight change, we can't fight gravity, we can't fight nothin'. My whole life, all I ever did was fight. But I can't give up neither, John, I can't fight my own nature. That's the paradox, John, you see? When I'm gone, they'll have to find a new monster. Our time is passed, John."
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For me, I think it'd be this^ song. The song itself combined with what's going on while the song plays... It was the only time throughout Twilight Princess or any other modern Zelda game really, where I actually thought that a character besides the villain might die.