The killings rocked the country, hastened the passage of the Civil Rights Act and were later dramatised in the film Mississippi Burning.
Killen was pronounced dead at 9 p.m. Thursday in the Mississippi State Penitentiary hospital, prison authorities said.
Killen's conviction came 41 years after James Chaney, Michael Schwerner and Andrew Goodman, all in their 20s, were ambushed and killed by Klansmen.
Their bodies were found 44 days later, buried in an earthen dam, after an extensive FBI investigation. The dramatic search for the missing men drew national attention to the violent resistance to the civil rights movement.
His first trial in 1967 ended in mistrial, but Killen was retried almost 40 years later after state authorities reopened the murder investigations, according to the Clarion Ledger. The state of MS didn't press charges at the time.
On June 21,1964, the three - all civil rights workers registering African-Americans to vote - were on their way to investigate the burning of a black church in Neshoba County when a sheriff's deputy took them into custody for speeding.
Killen was the only person charged, even though other suspects were still living.
Edgar Ray Killen, 92, was jailed 2005, over four decades after he plotted the killings.
Eighth District Attorney Mark Duncan, now a circuit judge; Mississippi Attorney General Jim Hood; Special Assistant Attorney General Lee Martin and others prosecuted the case.
In 2005, a Neshoba County grand jury indicted Killen for murder in the slayings of Chaney, Goodman and Schwerner.
The jury convicted Killen of manslaughter, which some of the victims' families found insufficient.
The cause death is pending an autopsy.
In 1999, MS authorities reopened the case after The Clarion Ledger reported the contents of a secret interview that Bowers had given in which he said he was "quite delighted to be convicted and have the main instigator walk out of the courtroom a free man".
At age 80, he was convicted on June 21, 2005, based on new evidence unveiled in 2000 and was sentenced to three consecutive 20-year sentences.