La Liga champions Real Madrid suffered a dent to their title defence with a 2-1 defeat at Real Betis on Saturday.
Isco scored the winner from the penalty spot against his former side, who are level on points with league leaders Barcelona, who face Real Sociedad on Sunday.
Atletico Madrid, third, host Athletic Bilbao later Saturday and can move top with a victory.
Real Madrid coach Carlo Ancelotti picked a strong line up, with no breathing room in the title race, despite the impending Champions League clash against city rivals Atletico.
“If we play like this on Tuesday (against Atletico) we will not win,” said Ancelotti.
The visitors took the lead at the Benito Villamarin stadium through Brahim Diaz but Betis struck back with a Johnny Cardoso header, as Real Madrid missed the suspended Jude Bellingham.
Isco secured the Andalusian side a deserved victory with a penalty early in the second half.
“It’s a big blow. We have to react,” continued Ancelotti.
“We started well, but later we did not have the same level of commitment, the game got out of control against a side that played better than us and deserved to win.”
Isco returned to action in December after missing several months with a broken leg and has hit top form in recent weeks.
“I’m dead… it takes so much work to win games, against Madrid, who have so many options, it’s very hard,” an exhausted Isco told Movistar.
“About halfway through the first half we stepped up our game, and I’m happy with the win against the best team around these days.”
The 32-year-old playmaker said he always has “Madrid in my heart” after winning several major trophies with the Spanish giants before departing in 2022.
Mbappe subbed
Madrid started the stronger and opened the scoring after just 10 minutes, putting together a fine team move with 19 passes before Diaz tucked home.
Kylian Mbappe sliced open the Betis defence with a ball to Ferland Mendy in front of goal and the defender squared for Diaz to convert.
Betis found their footing and neither team were able to create danger until the hosts levelled midway through the first half.
Cardoso powered a header past Thibaut Courtois from Isco’s corner after escaping the attention of Madrid’s defence.
Manchester United winger Antony, on loan at Betis, was sent off last weekend but his red card was revoked enabling him to face Madrid and he was increasingly involved.
Courtois pulled off a superb save after a ball bounced towards his goal off the back of Aurelien Tchouameni.
Madrid began to lose the ball frequently and without Bellingham’s dynamism in midfield were outfought by Betis.
Manuel Pellegrini’s side were in the ascendency and took the lead early in the second half from the penalty spot when Antonio Rudiger crashed into Jesus Rodriguez as he ran in on goal.
Isco, enjoying a renaissance at Betis, smashed home from the spot against his former side for his sixth goal across all competitions.
Tchouameni was fortunate to get away with clipping Ricardo Rodriguez inside Madrid’s area and Cucho Hernandez dragged a shot wide of Courtois’ far post when well placed.
For the final 15 minutes, Ancelotti replaced Mbappe, who had dental trouble ahead of the game, with Brazilian teenager Endrick who had scored in the Copa del Rey in midweek.
“He had that (tooth) problem, he hadn’t trained a lot,” explained Ancelotti.
“He was not at his best level and to avoid problems I took him off and put on Endrick, who has been doing well.”
Endrick fired wide and Betis goalkeeper Adrian kept out a stinging drive from range by Vinicius as Madrid came up short.
