The music legends teamed up for their own interpretation of the traditional hymn 'Will The Circle Be Unbroken', which dates back to 1935.
The pair last performed together at New York's Roseland Ballroom in 1994.
The 'Desolation Row' hitmaker and the former Crosby, Stills & Nash star both have a history with the song.
Dylan played the track on his tour of coffee shops in 1961, and six years later he record it for his 16th studio album 'The Basement Tapes' with The Band, which was released in 1975.
The 78-year-old musician hadn't included the song in his set since May 1976, when he performed in Houston, Texas, on his 'Rolling Thunder Review Tour'.
Young has performed the song four times since 1995 with Willie Nelson.
However, his first live performance of the hymn was in 1975 at the Kezar Stadium in San Francisco for a benefit show, where he was joined by Dylan for their first duet together.
The Kilkenny concert followed the pair's sold-out show in Hyde Park last week.
The two stars came together for an historic night in London which saw them play a host of their classic tracks for the capacity crowd of 65,000.
First on stage was Young who joked that he had ''never played in daylight before'' in reference to the fact he had to go on before Dylan and the Canadian musician was in good spirits as he added ''It's great to see everybody''.
Backed by his band The Promise of the Real, which features Willie's son Lukas, the rocker opened with 'Mansion on the Hill, 'Over and Over' and 'Country Home' from his Crazy Horse period before playing 'Alabama' - which features on his acclaimed 1972 album 'Harvest' - to huge cheers from the crowd.
Those cheers only got louder when Young strapped on his acoustic guitar and harmonica for beloved classic 'Heart of Gold', also taken from 'Harvest'.
Appreciating the crowd's response to the song, he followed it up with 'From Hank to Hendrix' and then acknowledging his fans' love of his folk material he said with smile whilst pointing to his guitar ''Do you want another one on this one?''
The huge cheers gave way to a rendition of 'Old Man' which sparked the biggest singalong of his set.
Young - whose stage was adorned with a Love sign and a bird flying above the band with a peace sign as an eye - also delivered an impassioned version of 'Rockin' in the Free World' showing his political passions still burn as brightly as ever.
He and the band returned for an encore of fan favourite 'Like A Hurricane' and 'Piece of Crap' - from his 1994 album 'Sleeps with Angels'.
It was then Dylan's turn to take to the stage and he walked out unannounced and sat down at his piano with a wide grin before opening with 'Ballad of a Thin Man' from his seminal 1965 album 'Highway 61 Revisited'.
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