Cuba's crackdown brought protests from families of the detainees
The United States has ordered 14 Cuban diplomats serving at the United Nations and in Washington to leave the country, implying that they had been spying.
The Cubans were being expelled "for engaging in activities deemed harmful to the United States" - the usual diplomatic way of referring to espionage.
Havana is likely to see the expulsions as a sharp escalation in the confrontation between the two nations, the BBC's Greg Barrow at the UN says.
Relations between Cuba and the US, poor at the best of times, have recently deteriorated significantly.
The US was highly critical when Cuba imprisoned 75 journalists, librarians and opposition figures and executed three Cuban hijackers who tried to take a boat to the US.
The executions triggered international protests, including condemnation by the Pope.
The Cuba Policy Foundation, a lobbying group dedicated to ending the US embargo of Cuba, disbanded in protest at the crackdown on dissent.
But more than 160 intellectuals and artists - including Nobel Prize-winning author Gabriel Garcia Marquez - signed a declaration in support of Cuba that was read out in Havana on May Day.
Garcia Marquez expressed support for Cuba
The statement condemned what it called the "harassment of Cuba".
US diplomats walked out of a UN meeting last month when Cuba was re-elected unopposed to the UN Human Rights Commission.
Cuba, meanwhile, has ordered American diplomats not to travel outside Havana without getting prior approval.
The government in Havana has portrayed its clampdown as a response to an allegedly power-crazed US President George W Bush, who, it says, might launch an attack on the island at any time.
"We do not want the blood of Cubans and Americans to be shed in a war," Cuban President Fidel Castro said on 1 May.
The US has not named the diplomats to be expelled or characterised their ranks. They were ordered to leave within 10 days of the delivery of a diplomatic note on Tuesday morning, the US State Department said.
They include seven from the UN and seven from Washington - but not Dagoberto Rodriguez, the head of the Cuban Interest section and highest ranking Cuban diplomat in the country.
Cuba is authorised to have up to 26 diplomats at its interest section and 37 at the UN. There are 51 US diplomats in Havana, the State Department said.
Tuesday's announcement of the expulsions comes as the White House prepares to announce possible changes to its Cuba policy.
The US often reveals new plans on 20 May, the anniversary of Cuban independence.