The negotiating teams met at the State Department just before 9:00 am (1400 GMT) for round two, after an initial meeting in Havana last month broke the ice but ended with little sign of a breakthrough.
The talks are building on US President Barack Obama and Cuban president Raul Castro's surprise announcement in December that the two Cold War adversaries had decided to normalise relations severed in 1961.
The US side in the talks is represented by Roberta Jacobson, assistant secretary of state for Western Hemisphere affairs, who faced Josefina Vidal, the Cuban foreign ministry's director for US affairs, across a long table flanked by their respective delegations.
A senior State Department official said the negotiators would review provisions in the Vienna Convention on Diplomatic Relations on such subjects as the freedom of movement of diplomats and the use of diplomatic pouches.
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The hope is that within the coming months both nations will agree to reopen embassies in each other's capitals and appoint fully-fledged ambassadors. Currently they operate with so-called interests sections in Havana and Washington.
US President Barack Obama is due to attend the Summit of the Americas in Panama on April 10-11, which Cuba will also attend for the first time.
But after more than five decades of hostility and suspicion, steep obstacles remain to renewing diplomatic ties.
This is seen as the first step towards a full normalisation of relations between the United States and the communist-run Caribbean island, which has been governed since a 1959 revolution by revolutionary leader Fidel Castro and now by his brother Raul.