Germany international Marco Reus has failed to make Joachim Low's final 23-man squad for Euro 2016, but Bastian Schweinsteiger and Mats Hummels are included.
Borussia Dortmund forward Reus, who missed the 2014 World Cup through injury, has been struggling with a groin problem and the 27-year-old will not travel to France.
"Reus can only run straight right now," Low told a news conference on Tuesday. "He could do no more in training. The medical staff was sceptical that he would be fully fit.
"It's a bitter decision for him and us. He would be a major asset for our team if he was fit."
Dortmund later confirmed on their official website that Reus currently has an inflammation of the adductor region.
Sami Khedira, who captained Germany in the 3-1 loss against Slovakia on Sunday, added: "It's a bitter blow for him. Just like 2014, it's a major setback for him. I am convinced he'll get another chance on the biggest stage."
Bayer Leverkusen duo Karim Bellarabi and Julian Brandt and Hoffenheim's Sebastian Rudy have also been cut from the 27-man provisional squad that Low named on May 17.
That means captain Schweinsteiger and defender Hummels make the squad despite concerns over their fitness.
Schweinsteiger, 31, partially tore the medial collateral ligament in his right knee while with the national team in late March but said at the start of this month that he was "very confident" he would be fit for the tournament.
Hummels, 27, suffered a torn calf muscle in the DFB Pokal final on May 21 and has said he faces a race against time to be fit for Germany's opening match against Ukraine on June 12.
"Hummels and Schweinsteiger will be available during the tournament," Low added. "Schweinsteiger will resume training today, Mats needs a few more days."
Low, who oversaw Germany's triumph at the 2014 World Cup, said it had been tough to make the final omissions.
"We've discussed the decisions we made yesterday and the day before yesterday," he said. "We talked to the medical staff. Those decisions hurt me on a human level. It's a big disappointment for the players. They have to look at it as an opportunity and as motivation."
Germany squad for Euro 2016
Goalkeepers: Manuel Neuer (Bayern Munich), Bernd Leno (Bayer Leverkusen), Marc-Andre ter Stegen (Barcelona)
Defenders: Jerome Boateng (Bayern Munich), Mats Hummels (Borussia Dortmund), Emre Can (Liverpool), Jonas Hector (Cologne), Benedikt Howedes (Schalke), Shkodran Mustafi (Valencia), Antonio Rudiger (Roma)
Midfielders: Mario Gotze (Bayern Munich), Mesut Ozil (Arsenal), Sami Khedira (Juventus), Toni Kroos (Real Madrid), Julian Draxler (Wolfsburg), Joshua Kimmich (Bayern Munich), Bastian Schweinsteiger (Manchester United), Julian Weigl (Borussia Dortmund)
Forwards: Thomas Muller (Bayern Munich), Andre Schurrle (Wolfsburg), Lukas Podolski (Galatasaray), Mario Gomez (Besiktas), Leroy Sane (Schalke)