Fred Astaire, and our friend Windypundit, have said it was the Nicholas Brothers in Stormy Weather. I beg to differ.
It was the Berry Brothers, perhaps not as technically perfect as the Nicholas Brothers (with whom they were rivals), but far more athletic, in the relatively obscure film Panama Hattie. Feast your eyes and ears on this.
They're like Jedi in tophats, tails, and canes.