The four-minute spot was directed and co-written by Barry Levinson, the man behind films such as 'Rain Man' and 'Good Morning Vietnam', and features an animated Superman alongside Seinfeld.
The new ad appears on the American Express website, along with a spot that first appeared in March this year.
Called "Hindsight is 20/20", the new ads shows Seinfeld and Superman going on a road trip. The pair stop to admire the landscape and take a picture when Superman accidentally locks the keys in Seinfeld's vintage Porsche.
At first Superman offers to rip one of the doors off to retrieve the keys, or to fly off and get help, but Seinfeld overrules, saying "It's a road trip, no flying." He then remembers that as a member of American Express he is eligible for roadside assistance, and soon the pair are on their way again.
The ad follows on from an earlier "webisode" called "A Uniform Used to Mean Something", where Superman and Seinfeld are walking through Times Square after a visit to a diner. That spot promoted American Express' guarantee to replace damaged goods within 90 days of purchase and first appeared in March.
After viewing the spot, users are asked if they wish to apply for an American Express card. The site also contains behind-the-scenes footage and clips from teasers running on television, to encourage people to look at the site.
American Express has a long-running relationship with Seinfeld and Superman, who last appeared together in an ad for the credit card in 1998.
Seinfeld, a long-time fan of the Man of Steel, said: "Our exploits make for great material, and I think the internet provides the perfect platform to share our comical adventures with fans around the planet."
John Hayes, American Express’ chief marketing officer, said of the campaign: "We are continually looking for innovative ways to engage consumers and introduce new customers to the brand, and the internet has already proven to be an exciting channel for us."
If you have an opinion on this or any other issue raised on Brand Republic, join the debate in the Forum here.