The 28-year-old singer joined Lambert and her rock band idols on stage to sing "Another One Bites the Dust" in Sydney. Lambert is fronting Queen for a summer tour, and has been on the road with the band since June 19.
Queen inspired Lady Gaga's stage name, and the singer posted a starstruck tweet and pictures with Lambert and the band after the performance.
"I just played with Queen I'm the happiest girl alive #QAL," she tweeted. "Tonight I have felt alive in a way I have not before. I returned to the Radio GaGa mothership to pay homage to my leaders. It was emotional and wild," she added on Instagram.
Lady Gaga will bring her ArtRave: The Artpop Ball tour to Sydney on Saturday. The singer released Artpop in November 2013, and will close the tour in Paris on November 24.