Having both failed to win last time out, the sides will be looking to round off what has been an otherwise disastrous continental campaign on a high note.
Match preview
Defensa were held to a 2-2 draw by Ecuadorian outfit LDU Quito in Thursday's meeting at the Estadio Rodrigo Paz Delgado.
Sebastian Beccacece's men went behind to Jose Quinteros's early opener before a quickfire double from Lucas Albertengo turned the game on its head, only for Nilson Angulo to restore parity 19 minutes from time.
As a result, The Halcon now sit third in Group C, a whopping eight points behind group leaders Atletico Goianiense with just one game left to play.
The Buenos Aires-based side now set out to claim a second straight win over their next opponents, who fell to a 3-1 defeat in last month's reverse fixture.
Off the back of recently ending five-game home winless streak, Wednesday's hosts will be looking to record consecutive victories in front of their fans for the first time since November.
Antofagasta were beaten 1-0 by Atletico Goianiense on Wednesday when the teams went head to head at the Stadium Antonio Accioly.
Jorge Jorginho's 12-minute strike proved to be the difference as Juan Manuel Azconzabal's men failed to score for the third game on the spin.
However, The Pumas put an end to that dismal run in front of goal yesterday following a 2-2 draw versus Everton in the Chilean Premier Division.
Leading up to Wednesday's contest, the visitors are on a five-match losing run on the road with their last away triumph coming back in March.
Defensa y Justicia Copa Sudamericana form:
Defensa y Justicia form (all competitions):
Antofagasta Copa Sudamericana form:
Antofagasta form (all competitions):
Team News
With four goals on his last four matches across all competitions, Defensa's Lucas Albertengo is definitely one to look out for.
Miguel Merentiel, who put on a superb attacking display in the reverse fixture, is expected to return to the starting XI for this one.
Having gone five games without a clean sheet, Juan Manuel Azconzabal could opt to make some alterations to his regular defensive unit.
This could see Paulo Magalhaes and Rojas Sanhueza get the nod to feature in place of Salvador Cordero and Robles Fuentes.
Defensa y Justicia possible starting lineup:
Unsain; Soto, Frias, Colombo, Tripichio; Hachen, Gutierrez; Rotondi, Bou, Pizzini; Merentiel
Antofagasta possible starting lineup:
Gonzalez; Magalhaes, Vega, Rojas, Rodriguez; Hurtado, Orellana, Bravo, Flores; Lopez, Ramos