The item was described in the online listing as being a 4.5-inch "mini bible" signed by the president at a rally in 2016, which would have been before his inauguration.
Justin Steffman, the CEO and lead authenticator of AutographCOA.com, examined photos of the Bible and its signature and told NBC News that the autograph was counterfeit.
"I have taken a close look and compared the book to our authentic exemplar database," Steffman said by email.
"The signature on the Bible definitely does not appear to be an authentic signature from President Trump. The odds of it being real are zero per cent in my opinion."
His opinion was backed by two authenticators from the firm PSA/DNA, Kevin Keating and Tom Poon, who separately came to the same conclusion.
"Both of them came back almost immediately with thumbs down," said Terry Melia, spokesman for the company.
Melia added: "The percentage of fake autographs being sold on eBay — ones that do not already include third-party authentication behind them — is easily 50 per cent or more."
A certificate of authenticity was provided with the Bible that had allegedly been signed by Mr Trump.
The seller has responded to NBC News' questions about the eBay listing: "Do not contact me again or I will contact my lawyer and get you for harassment."
Ryan Moore, an eBay spokesman, said by email, "Our policies require that items listed on eBay are authentic.
"In the rare case a buyer believes they have purchased an item that is inauthentic, eBay's Money Back Guarantee will cover them."
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