Aguero has been out of action since suffering a knee injury during a Premier League victory over Burnley in June.
The Argentine recently stepped up his recovery in the gym after undergoing surgery on the issue, and Guardiola dropped the biggest hint yet over a possible return date for the striker prior to City's final day win against Norwich at the weekend.
"I think against Madrid everyone will be fit," the 49-year-old head coach said before watching his side beat the Canaries 5-0 at the Etihad Stadium.
After the match, however, Guardiola performed a U-turn regarding the status of Aguero, who was left on the bench when City beat Madrid 2-1 in the first leg of their round of 16 tie at Santiago Bernabeu back in February.
When asked if the 32-year-old will be a part of his squad for the second leg on August 7, the City manager told reporters: “No, I think it will not be possible for the timing.”
Aguero has 23 goals to his name from 32 appearances in all competitions this season, with Raheem Sterling the only man in the City squad currently boasting a higher tally, as he reached the 30-mark for the first time in his career by scoring against Norwich.
Gabriel Jesus will likely continue to lead the line in Aguero's absence when Madrid arrive at the Etihad, having scored an all-important equaliser at the Bernabeu earlier this year.
City's latest five-goal haul in the Premier League took them to 100-plus goals in the competition for the third time in their history, but Guardiola insists that thrashing Norwich will have little bearing on the Madrid encounter.
“Different competition, different opponent, cannot compare,” he added. “We have two weeks.
"We are going to start to prepare the game, tell them what they should do, try to play the game, do a good game. Arrive in the best conditions and we are going to do it.”