Neville has been out of action for the club since breaking an ankle in March but was was expected to feature after making an appearance in a reserve team game against Stockport County last Thursday.
However, Sir Alex Ferguson has revealed the 32-year-old is now back on the treatment table after suffering the injury in training.
The United manager said: "Unfortunately, Gary has a little calf strain."
“It’s really bad news for the boy. I don’t think it is too bad but it is one he could have done without.
“He needs to get his season on the road and this is a setback.”
Not only will the Manchester United right-back miss out on helping his team secure a place in the knock-out stage, he is also likely to be sidelined for upcoming England internationals.
Steve McClaren is due to name his squad on Friday for this month's friendly against Austria and the Euro 2008 qualifier with Croatia.