Isco has joined Real Madrid's mounting injury list after suffering a hamstring issue.
Scans on Wednesday revealed muscle damage in the attacking midfielder's right leg.
Spain international Isco was substituted in the 68th minute of the weekend meeting with Real Valladolid as Madrid was held to a 1-1 home draw .
The Spanish side has not confirmed the severity of the setback, but the 27-year-old is likely to miss Sunday's visit to Villarreal.
"His recovery will continue to be monitored," the club added in a statement.
Isco's injury is another blow to Los Blancos boss Zinedine Zidane, who has been without Eden Hazard, Ferland Mendy and long-term absentee Marco Asensio during La Liga's opening two rounds.
Los Blancos have also lost James Rodriguez to a calf problem, while youngsters Rodrygo Goes and Brahim Diaz remain sidelined.
Isco might additionally be forced to sit out his country's Euro 2020 qualifiers against Romania and Faroe Islands on September 5 and 8, respectively.