It was noted for its holding the high ground at the center of the line at Antietam as part of Tyndale's 1st Brigade, Greene's 2nd Division of Mansfield's XII Corps.

Pushing the Confederates south to the Dunker Church, Tyndale's men held the area until the afternoon when lack of support, heavy losses and low ammunition compelled them to retreat.

[4] In 1863, the partially replenished 5th Ohio Infantry fought in the Army of the Potomac at Chancellorsville and then served in the Gettysburg Campaign.

Transferred later in the summer to the Western Theater and becoming part of the XX Corps under Joseph Hooker, the regiment participated in the Battle of Lookout Mountain near Chattanooga, Tennessee.

During the Civil War, the 5th Ohio Infantry participated in 28 battles and sustained a loss of more than 500 men killed, wounded and prisoners.