According to Spanish media, the federation’s Appeals Committee said it was not overturning the two-game suspension handed to Neymar for sarcastically applauding an assistant referee in Barcelona’s 2-0 loss at Malaga on April 8.
The two-match ban was in addition to a one-game suspension he was automatically required to serve after being sent off in Malaga.
Neymar served that suspension last weekend as Barcelona beat Real Sociedad 3-2.
In addition to El Clasico on Sunday, Neymar will also miss the Spanish league game against bottom -placed Osasuna on April 26.
Barcelona trail Real Madrid by three points with six rounds to go in the LaLiga Santander.
In the first El Clasico, this season at the Nou Camp Real Madrid forced Barcelona to a 1-1 draw.