A portrait widely thought to be William Shakespeare is not the playwright, the National Portrait Gallery has revealed.
Technical analysis has confirmed its date and English origin
The Grafton Portrait has been used on the covers of many books on the Bard.
But nine months of research and restoration work have failed to reveal any evidence to support the view that it is a portrait of Shakespeare.
The gallery said it dates back to 1588 when Shakespeare was 24, but at that stage he was unlikely to have been able to afford the style of dress seen.
Gallery experts said at that age he was about to join a travelling theatre troupe and had just become a father to twins.
It follows confirmation in April that The Flower Portrait, owned by the Royal Shakespeare Company, was a fake.
The Grafton Portrait was painted in oils by an anonymous artist and shows a youth with curly brown hair and grey eyes, and is dressed in a sumptuous silk or satin scarlet jacket worn by European men between the 15th and 17th centuries.
The inscription on the top of the painting records the age of 24 and the date as 1588, which would make the sitter the same age as Shakespeare.
Manchester's John Rylands Library has loaned the portrait to the gallery for restoration ahead of its Searching for Shakespeare exhibition.
'Shakespeare in Love'
Dr Tarnya Cooper, 16th Century curator at the National Portrait Gallery, said: "We believe that Shakespeare left Stratford-upon-Avon following the birth of twins in 1585.
"One possibility is that he joined a travelling theatre troupe and it is very unlikely that in 1588, Shakespeare would have been able to afford a costume of this type."
She said the painting has not been looked at in a systematic way before.
"But the painting has fuelled the kind of Shakespeare in Love theories of the 21st Century, of a beautiful young man with a sensitive and passionate face, of a character with an incredible emotional range," she said.
The so-called Chandos portrait, which has been the subject of fierce debate over whether it really features Shakespeare, is next in line for examination.
The Culture Show broadcasts a report into this issue on BBC Two on Saturday at 2315 BST.