Spain coach Vicente del Bosque has hinted that David de Gea is likely to start in goal.
Striker Alvaro Morata is fit after missing their warm-up match against Georgia with a thigh injury, while David Silva could win his 100th cap.
Czech Republic's Borek Dockal has been hampered by an ankle injury, but is expected to play.
Marek Suchy is suspended for the opener so Michal Kadlec or Roman Hubnik will replace him at centre-back.
Spain lost their final warm-up match to Georgia - ranked 137 in the world - on Tuesday. Three days later goalkeeper David de Gea denied a claim reported in a Spanish newspaper that he organised a sex party with prostitutes for teammates in which a woman was made to take part against her will. Up until then, the two-time defending champions had been quietly approaching Euro 2016 under the radar.
Which is odd for a side that have not lost a match at a European Championship finals since 20th June 2004, and whose last defeat in a competitive fixture was back in October 2014.
Opponents Czech Republic also suffered a defeat in their final warm-up match. They lost 2-1 to a South Korea side that had been routed 6-1 just days earlier… by Spain. In fact, Pavel Vrba's side have won just one of their last six matches and have a real difficulty keeping clean sheets.
Spain's attackers must be licking their lips in anticipation of facing a side who failed to record a shut-out in qualifying, and conceded the most goals (14) of all the teams to make it to Euro 2016.
Spain are unbeaten in four games against the Czech Republic. The Czechs have scored only one goal in those four matched (netted by Jaroslav Plasil in March 2011).
Spain qualified for the 1998 World Cup at the Czechs' expense. The only goal in their two qualifiers was a penalty converted by Fernando Hierro.
The teams also met in Euro 2012 qualifying. David Villa scored twice for Spain in a 2-1 win in Granada, with Juan Mata and Xabi Alonso on target in their 2-0 away victory.
Spain won four of 12 encounters with the former Czechoslovakia (D1, L7). In competitive games, the Spanish won two and lost three of five meetings.