Barber was born to Thomas and Mary Oliver Barber in Franklin County, Pennsylvania. In the early 1830s, he moved to Richmond, Indiana, where he was engaged for some time in operating a woolen mill. Soon after the passage of the Kansas-Nebraska Act, he moved to Kansas and settled on a claim some seven miles southwest of Lawrence. Being a sober, honest, and industrious, he made friends among his neighbors. Early in December 1855, when the pro-slavery forces were threatening Lawrence, Barber decided to go to the town’s assistance. He had no children, but his wife, who seemed to have had a premonition of impending danger, begged him to remain at home, but he laughed at her fears and set out on horseback for Lawrence. On the morning of December 6, 1855, in the company of his brother Robert and a man named Thomas M. Pierson, he started for his home, unarmed, promising to return as soon as he had arranged matters to permit his absence.
About four miles from Lawrence, on the California Trail, they saw a party of 14 horsemen approaching; two of them rode on in advance of the others to converse with Barber and his companions. These two men were George W. Clark, an agent of the Pottawatomie Indians, and a merchant of Weston, Missouri, by the name of Burns. They tried to induce the Barbers and Pierson to join them, and meeting with a positive refusal, one of them drew his revolver and fired twice, mortally wounding Thomas W. Barber. He concealed that he was shot until they had ridden about 100 yards when he informed his brother, who initially thought such a thing impossible, but a few minutes later, the wounded man was seen to reel in his saddle. His associates eased him to the ground, where, a little later, he inhaled his last breath on December 6, 1855. The 41-year-old hero was buried in the Pioneer Cemetery in Lawrence, Kansas.