The third-tier Cipolletti upset a side in the division above them – Villa San Carlos – 4-2 on penalties after their clash finished locked at 1-1.
Santiago Vergara had given visitors Cipolletti a 77th-minute lead with a spot-kick, but his side were reduced to 10 men moments later when Lucas Mellado saw red.
Substitute Diego Mendoza struck from the penalty spot for the hosts five minutes later and a shootout followed.
And Cipolletti got the better of their more-fancied rivals in the spot-kicks to claim an upset victory.
Elsewhere, second-tier Brown de Adrogue battled past fourth-tier Villa Cubas 3-2.
Juan Manuel Garcia netted a brace for the winners, who were never behind and held on at the Estadio Bicentenario Ciudad de Catamarca.
Second-tier Instituto got past Chacarita Juniors 3-1 and Crucero del Norte were 2-0 victors at home to Gimnasia Concepcion thanks to Victor Aquino's brace.
‘After Manchester City’s recent form, maybe they’re the underdogs against Manchester United!’ Former Red Devils defender on this weekend’s derby
‘Arteta, Alonso, Emery, me… none of us were physical players – we needed the understanding of the game. That probably helped us move into management’: Premier League boss reveals reasons for natural career progression
