Lister, the son of Joseph Jackson Lister, was born at Upton in England and educated at Quaker schools before entering University College, London, in 1843. University College was, at the time, the only English university open to religious dissenters. After graduating in arts, Lister studied medicine, obtaining his MB in 1852. He then served as assistant to the leading Scottish surgeon James Syme at the Royal Infirmary, Edinburgh, from 1854 until 1860 when he was appointed professor of surgery at Glasgow University. Lister returned to Edinburgh in 1869 as professor of clinical surgery but in 1877 became professor of surgery at King's College, London, serving there until his retirement in 1892.

In 1867 Lister published two short but revolutionary papers, which introduced the principles of antiseptic surgery into medicine. In 1846 he had been present when Robert Liston had first successfully used ether as an anesthetic in England. Yet this, apart from making surgery tolerable for both patient and surgeon, had not greatly advanced the profession. The full potential of anesthesia did not develop because of the high mortality produced by the infection that inevitably followed major surgery. The inevitable consequence was that a surprisingly small amount of surgery was actually attempted, even in the major centers with ready access to anesthetics.

Lister acknowledged the twin sources of his innovations in his 1867 papers. The first and most important were the writings of Louis Pasteur. These revealed the cause of the widespread surgical sepsis to be the germs present in the air. To control them Lister reported that he had been impressed by an account of the effects produced by carbolic acid on sewage in Carlisle. Carbolic acid (phenol, C₆H₅OH) is a weak acid derived from benzene. Although Lister's first attempt to use it as an antiseptic in March 1865 ended in failure he persisted and in August dressed a compound fracture of the leg, that is one in which the skin has been broken, with a piece of lint dipped in liquid carbolic acid. The wound healed well.

This encouraged Lister to introduce the carbolic acid dressings into his regular surgical procedure. By 1870 he claimed that mortality for amputations had dropped from over 40% to 15%. He later analyzed his figures for his Edinburgh period, reporting that from 1871 to 1877 he performed 725 major operations with a mortality of only 5.1%.

Another of Lister's major innovations was his introduction, in 1869, of cat-gut ligatures to replace the traditional silk thread, which was a major source of infection. Lister's experiments showed that cat-gut ligatures were absorbed by the body and if soaked in carbolic acid could be made sterile. He also attempted to maintain an antiseptic atmosphere in the operating theater by introducing a carbolic spray. This, however, made working conditions very unpleasant and the procedure was abandoned.

The Listerian system appears to have been accepted with little dissent and remarkable speed for by 1880 it had become the standard mode of surgical procedure virtually everywhere. Lister, an intensely shy and reserved man, achieved considerable fame and received many honors. In 1897 he became the first physician to be made a peer and sit in the House of Lords.