The page contains the original features of the hero's black symbiote costume that later led to the creation of anti-hero Venom.
A single page of original artwork from an acclaimed 1984 comic featuring Spider-Man sold for a record $3.36 million in the United States.
The record price made the webslinger the world's mightiest auction superhero on Thursday.
The page features the first appearance of Spidey's black symbiote suit that would later lead to the creation of anti-hero Venom in artwork by Mike Zeck from Marvel Super Heroes Secret Wars no. 8.
"This page was the big reveal teased on the cover! This is where Peter Parker actually got his spiffy new black costume," Heritage Auctions said in its description of the artwork for page 25 of the issue.
"But... it's a costume with a secret! Because it very soon turns out to be alive and have its own agenda. This is the origin of the character Venom!"
Bidding for the artwork had started at $330,000.
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#SpiderMan's Black Costume Origin Sells for $3.36 Million at Heritage Auctions to Shatter #ComicArt Record.#Superman also breaks $3 million barrier with Action Comics No. 1 sale to kick off four-day #Comics and Comic Art event. 💥https://t.co/MPvQamZcei#MarvelComics pic.twitter.com/iB7RyRsAeF— Heritage Auctions (@HeritageAuction) January 13, 2022
The previous record for a single page of artwork from the interior of an American comic book was a frame showing the first image of Wolverine in a 1974 issue of "The Incredible Hulk". The page sold for $657,250.
A copy of 1938's Action Comics no. 1, the first appearance of Superman, sold for $3.18 million on the first day of Heritage Auctions' four-day comic event in Dallas, Texas, the auction house said on Twitter.
The copy was graded 6.0 by comic book grading authority CGC.
Two other copies of the world's most famous comic book issue with a higher grade have previously sold for more at auction, according to The Hollywood Reporter.