England's Frank Lampard will undergo a thigh scan on Thursday after pulling up in training ahead of Saturday's pre-Euro 2012 friendly against Belgium.
The 33-year-old Chelsea midfielder will be assessed by England's medical team.
Liverpool's Jordan Henderson, 21, has been put on standby as cover while Lampard is monitored.
Manager Roy Hodgson is already without Gareth Barry, who was ruled out of the tournament after suffering a groin injury in the 1-0 win over Norway.
Lampard, who has 90 caps, joined up with the England squad on Tuesday after playing in Chelsea's Champions League final win over Bayern Munich on 19 May.
Along with team-mates John Terry, Ashley Cole and Gary Cahill, he was given time off to recover from the drama of the penalty shoot-out victory.
BBC Sport's chief football writer Phil McNulty said Lampard would be a big loss to England, if the injury rules him out of the tournament.
"Frank Lampard has occasionally been a maligned figure for England - but his loss from the Euro 2012 squad would be a heavy blow for manager Roy Hodgson.
"Lampard has maturity, experience and continues to be a winner as proved by his vital role in Chelsea's Champions League and FA Cup victories.
"He may not have started England's first Euro 2012 game against France but his presence and professionalism around the squad is always beneficial and he would have been determined to force his way into the side.
"Liverpool's Jordan Henderson is in his infancy as an England international so Hodgson will be desperate for an optimistic medical bulletin on Lampard's thigh injury," said McNulty.
Before his injury setback, Lampard had spoken to the media about his Euro 2012 preparations.
"We've all had a bit of time to be with our families," he said.
"It has given us a nice period of rest for not only our bodies but our minds."
With less expectation around England's chances than at major tournaments in recent years, Lampard is optimistic the national side can emulate Chelsea's "against the odds" triumph in the Champions League.
"At Chelsea we achieved something potentially against the odds - maybe England will be the same," he said. "I think we have got the quality, we have to believe we can go all the way."
England play France on 11 June in their first match in Group D, before facing Sweden (15 June) and tournament co-hosts Ukraine (19 June).
"I think it's a tough group," Lampard added. "When people first saw it they were probably pretty happy but, when it sinks in, there are a lot of steps to overcome to get through the group. I believe we have the ability to get through it, though."
England forward Theo Walcott expressed his hope the midfielder will be fit for the tournament and added: "When you have a player who has just won the Champions League it will be a boost to the players and give an extra incentive, but it's early days (with the injury) yet."
BBC Match of the Day presenter and former England striker Gary Lineker said on Twitter: "If Lampard joins (Jack) Wilshere and Barry on the crocked list, copious amounts of cotton wool will be required for (fit midfielders) Steven Gerrard and Scott Parker."