News Chopper 2 was over the scene and video showed the injured animal sprawled on the interstate, next to the HOV lane, just east of Columbia Drive.
"Allegedly the buffalo fell out of the back of the trailer in which it was being transported," said Department Of Transportation spokeswoman Monica Luck in an e-mail to Channel 2 Action News.
The injured animal tried several times to get up, but failed.
GDOT spokeswoman, Erica Fatima said police "had to put the buffalo down" due to its injuries.
It's unclear why the animal was being transported or where to.
All lanes of I-20 were blocked.
The incident comes just eight days after a zebra escaped from a circus in downtown Atlanta and ran loose on the downtown connector.
The zebra was captured and returned to the circus.
SLIDESHOW: Zebra Caught In Downtown Atlanta