The 25th Massachusetts was organized at Camp Lincoln in Worcester, Massachusetts, beginning September 1, 1861 and mustered in for a three-year enlistment on October 31, 1861, under the command of Colonel Edwin M. Upton.
The regiment was attached to Foster's 1st Brigade, Burnside's Expeditionary Corps, to April 1862.
2nd Brigade, 1st Division, XVIII Corps, Department of North Carolina, to June 1863.
2nd Brigade, United States Forces, Portsmouth, Virginia, to April 1864.
1st Brigade, 2nd Division, XVIII Corps, Department of Virginia and North Carolina, to September 1864.
3rd Brigade, 1st Division, XXIII Corps, Department of North Carolina, to July 1865.
Burnside's Expedition to Hatteras Inlet and Roanoke Island, North Carolina, January 7 – February 7, 1862.
Guard, picket, and outpost duty at New Bern until December 10.
Moved to Newport News October 16–18, and duty there until January 22, 1864.
Moved to Portsmouth January 22, 1864, and duty in the defenses of that city until April 26.
Butler's operations on south side of the James River and against Petersburg and Richmond May 4–28.
Moved to White House, then to Cold Harbor May 28 – June 1.
Moved to New Bern, North Carolina, September 4–10, and duty there until March 1865.