Experts have discovered that a portrait of Mary, Queen of Scots, is 200 years older than previously thought.
The painting's oval surround had been hidden with brown paint
The painting, belonging to the National Portrait Gallery in London, was thought to date from the 18th Century.
However, tree analysis has revealed the panel was felled in the 16th Century and the work dates from between 1560 and 1592.
This means it would have been painted during the monarch's lifetime or very soon after her execution for treason.
The majority of surviving portraits of Mary Stuart (1542-1587) either come from the Jacobean period, when her son James came to the English throne, or are later.
The small portrait depicts the Catholic queen after her return to Scotland following the death of her first husband King Francis II of France.
During this period she remarried and gave birth to her only child, who later was to become James VI of Scotland and James 1 of England.
Mary was forced to abdicate the Scottish throne in favour of her son in 1567.
After becoming the focus for the Catholic rebellion, she was imprisoned by Elizabeth I and later executed.
It is believed that the painting may have been created as an image for one of Mary's supporters, either as a symbol of loyalty during her incarceration or as an icon of Catholic martyrdom just after her death.
During the investigation, experts also discovered that it was originally surrounded by an oval background which was later painted over with dark brown paint.
They said the repainting may have taken place in the late 18th or early 19th Century to fit in with a set of other images of European kings and queens.
Dr Tarnya Cooper, curator of 16th Century paintings at the National Portrait Gallery, said it was an "exciting find".
"The picture has not been on display for a long time because it was thought to be a much later copy," she said.
"It very satisfying to find out that the picture is far more important than we previously thought."
The painting is now coming out of storage and going on display at the Tudor Galleries in London.