Its discovery by Philip Mould, an art dealer, has been described as "important and exciting" by Dr David Starkey, the historian and television presenter who has written a biography of Elizabeth's early years.
The picture by an unknown artist is only the second known portrait of Elizabeth as a young princess, Times Online reports.
Mr Mould, a leading portraiture dealer who has a gallery in London, will be offering the painting for sale for about £800,000 at The Grosvenor House Art & Antiques Fair, which opens in Park Lane, Mayfair, on Thursday.
When he bought it at auction from a private Spanish vendor, the portrait was obscured by three layers of overpainting. Artists from the 18th, 19th and 20th centuries had tried to touch up the picture by brightening the face, Telegraph.co.uk reports.
Mr Mould suspected that there was a painting hidden beneath because parts of the portrait that had not been overpainted contained colours consistent with 16th-century pigments. He also attempted to date the portrait by analysing the wooden panel and the chemical composition of the pigments. Both tests suggested a 16th-century portrait. The tree used to make the panel was felled as early as 1546, suggesting that the portrait was completed between 1547 and 1553, when Queen Mary began her reign and portraits of Elizabeth as a teenager would have served no political purpose.
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