The deafening roars and piercing whistles of the Bilbao fans have often given their team extra impetus in the claustrophobic, 40,000-seater arena known as the Cathedral and the Basques are right in the tie after holding Barcelona to a 0-0 draw in last month's first leg at the Nou Camp.
"Bilbao are a team that will create problems for us," Barcelona coach Pep Guardiola said after the league leaders ground out a 2-1 home win over promoted Levante on Sunday.
Barca, who would meet Getafe or second-division leaders Real Betis in the last eight, won 3-1 in Bilbao in La Liga in September, when midfielder Andres Iniesta was roundly booed after he was involved in the dismissal of Fernando Amorebieta.
Spain midfielder Iniesta has mostly been hailed as a hero by opposing fans for scoring the winning goal in July's World Cup final but that achievement cuts little ice with the fiercely independent Basques.
"Now we have a life or death match," Iniesta said after the Levante game.
Barca's arch-rivals Real Madrid have not won the domestic Cup since 1993 but virtually secured a quarter-final berth with an 8-0 demolition of Levante in the first leg.
Jose Mourinho's side, who play Levante on Thursday, look likely to go on and face city rivals and 2010 runners-up Atletico, who won their first leg at home to Espanyol 1-0.
Mourinho said Brazil playmaker Kaka, who had knee surgery after the World Cup and has not played this season, could get a run-out in Real's La Liga match at neighbours Getafe later on Monday.
"He won't start against Getafe and he is still nowhere near being able to," the Portuguese told a news conference. "He could give us 15 or 20 minutes of his super quality but no more."
In other last 16, second legs, Villarreal host Valencia on Thursday after the pair drew 0-0 at the Mestalla, Malaga are looking to overturn a 5-3 defeat by holders Sevilla and Almeria will attempt to defend a 4-3 lead at Real Mallorca.
