The 20-year old talent has agreed a pre-contract to join the Premier League club this winter but his work permit application for the UK has been denied.
Agudelo had a work permit hearing on Wednesday in which Potters Chief Executive Tony Scholes and manager Mark Hughes presented the club's case to an appeals panel in London.
The panel decided to reject the appeal, giving Stoke no right to appeal again, effectively ending their pursuit of Agudelo.
"We're bitterly disappointed that the panel rejected our appeal for a work permit for Juan," Scholes told Stoke's official website.
"The criteria by which the panel should make work permit application decisions are well established and have been in place for some years and, despite recent comments to the media and discussion in the media, that criteria has not changed.
"We are therefore left amazed that our application for a work permit for Juan has been rejected when you compare his talent and ability to players who have been granted a work permit on appeal in the past.
"Unfortunately, under the rules of the appeal panel system we were given no explanation why the application was rejected and so can only speculate as to the reasons why we have been unsuccessful."
After the decision, Agudelo was taking a pragmatic approach to the bad news via Twitter.
God has a plan I know it.— Juan Agudelo (@JuanAgudelo) November 20, 2013
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