John Stones (Manchester City)
Despite being on the bench against Schalke in the Champions League last midweek, manager Pep Guardiola made it clear that the centre-back is troubled by a groin injury and as a result omitted him from his FA Cup squad.
Fabian Delph (Manchester City)
Delph was replaced less than an hour into Manchester City's FA Cup quarter-final win at Swansea last weekend and withdrew from the squad with an unspecified issue.
Ruben Loftus-Cheek (Chelsea)
The midfielder came on as a second-half substitute as Chelsea lost to Everton on Sunday but withdrew the following day through an undisclosed issue.
Played the full 90 minutes as Manchester United bowed out of the FA Cup at Wolves, but a reoccurring complaint saw the left-back pull out on Monday.
Trent Alexander-Arnold (Liverpool)
Came to St George's Park out of respect to Southgate and in a bid to overcome a back injury picked up when Liverpool played 10 days earlier. Made progress but returned to his club on Wednesday.
Marcus Rashford (Manchester United)
Played the entirety of United's FA Cup loss at Molineux, scoring at the end, but has been unable to shake an ankle injury during individual sessions. Returned to United on Thursday.