The Bureau of Prisons confirmed Ames died on Monday.
Ames, a 31-year CIA veteran, admitted being paid $US2.5 million by Moscow for US secrets from 1985 until his arrest in 1994.
His disclosures included the identities of 10 Russian officials and one Eastern European who were spying for the United States or Great Britain, along with spy satellite operations, eavesdropping and general spy procedures.
His betrayals are blamed for the executions of Western agents working behind the Iron Curtain and were a major setback to the CIA during the Cold War.
He pleaded guilty without a trial to espionage and tax evasion and was sentenced to life in prison without parole.
Prosecutors said he deprived the US of valuable intelligence material for years.
He professed "profound shame and guilt" for "this betrayal of trust, done for the basest motives," money to pay debts.
But he downplayed the damage he caused, telling the court he did not believe he had "noticeably damaged" the United States or "noticeably aided" Moscow.
"These spy wars are a sideshow which have had no real impact on our significant security interests over the years," he told the court, questioning the value that leaders of any country derived from vast networks of human spies around the globe.
In a jailhouse interview with The Washington Post the day before he was sentenced, Ames said he was motivated to spy by "financial troubles, immediate and continuing".
Ames was working in the Soviet/Eastern European division at the CIA's headquarters in Langley, Virginia, when he first approached the KGB, according to an FBI history of the case.
He continued passing secrets to the Soviets while stationed in Rome for the CIA and after returning to Washington.
Meanwhile, the US intelligence community was frantically trying to figure out why so many agents were getting discovered by Moscow.
Ames's spying coincided with that of FBI agent Robert Hanssen, who was caught in 2001 and charged with taking $US1.4 million in cash and diamonds to sell secrets to Moscow.
He died in prison in 2023.
Ames' wife, Rosario, pleaded guilty to lesser espionage charges of assisting his spying and was sentenced to 63 months in prison.
