Von der Leyen said she had been in a meeting with an unnamed individual who had tested positive for the virus.
European Commission President Ursula von der Leyen has tested negative for COVID-19, her spokesman has said.
It came after the leader said on Monday morning she would self-isolate until Tuesday after coming into contact with someone who tested positive for the virus.
She "will nevertheless observe self-isolation until tomorrow end-of-day as foreseen by health safety regulations," chief spokesperson Eric Mamer wrote on Twitter.
Tuesday marks seven days from her contact with the case of COVID-19.
The EU chief said in her tweet that she had received a negative result last Thursday as well.
She is not experiencing any virus symptoms for the moment, a spokesperson had said, but she will not be able to attend the European Parliament plenary.
The leader was last week in Lisbon for meetings with top Portuguese officials during which she had contact with a confirmed case. She learned of the contact late Sunday night, Mamer said at a press conference.
The president's announcement came after the European leaders' summit which was postponed when the president of the European Council, Charles Michel, last month was forced to quarantine for a week when a security guard in his team tested positive for COVID-19.
Von der Leyen attended the summit when it went ahead last Thursday and Friday, but did not know at the time that she had been in contact with a positive case.
She was also set to take part in a pivotal week for Brexit negotiations, having this week said she would work with UK Prime Minister Boris Johnson to thrash out a trade deal.
European Commissioner for Innovation Mariya Gabriel also announced via Twitter on Monday that she was self-isolating pending test results after a member of her team tested positive for COVID-19.
"Myself and my whole team are self-isolating in line with public health protocols and working from home. I feel well and have no symptoms. Have a good week, stay safe!" she wrote.
Staffers at the Commission are tested twice a week on Monday and Thursday. There have been at least 159 confirmed positive cases at the Commission.
This article has been updated: a previous version said Von der Leyen "was set" to go to Portugal.
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