Born at Castle Street, Dublin, Ware was the eldest son of James Ware, who arrived in Ireland in 1588 as a secretary to the Lord Deputy of Ireland, FitzWilliam. His father was knighted by King James I, was elected M.P. for Mallow in 1613, and served as auditor-general for Ireland till his death in 1632, in which capacity he was succeeded by his son. James graduated from Trinity College Dublin in 1616, having received a good education in Latin and Greek. Becoming interested in Irish history, he began assembling a very fine collection of Irish manuscripts, and made transcriptions from works held in other collections, including that of his close friend James Ussher, Bishop of Meath.
Knighted in 1629, Sir James was elected Member of the Irish House of Commons (M.P.) for the University of Dublin in 1634. In 1638 he, with Sir Philip Perceval obtained the monopoly of granting licenses for the sale of ale and brandy.
Ware's first book, published in 1626, was Archiepisco Porum Cassiliensium & Tuamensium Vitae, followed by Caenobia Cistertientia Hiberniae and De Praesulibus Lageniae, both in 1631. In 1633 he published three edited works: Edmund Spenser's 'View of the State of Ireland'; Meredith Hanmer's 'History of Ireland'; and Edmund Campion's 'History of Ireland'. His book of 1639, De Scriptoribus Hiberniae was to be the last published for fifteen years, due to his involvement in Irish and British politics.
Ware was an ardent royalist during the 1640s, during the Irish Confederate Wars, which was part of the conflict known as the Wars of the Three Kingdoms, which consumed much of Ireland and Britain during the 1640s. He was a strong supporter of James Butler, 1st Duke of Ormonde. His activities led to his imprisonment in the Tower of London for a year; following his release he returned to Dublin, only to be taken prisoner and hostage on the city's surrender to Colonel Michael Jones in June 1647. He was expelled to England that year, returned, and was expelled again in 1649. He spent some eighteen months exiled in France before travelling to London where he spent most of the 1650s.
Ware's first new book since the 1630s was De Hibernia et Antiquitatibus eius Disquisitones, published in London in 1654, and in a second edition in 1658. This was followed in 1656 by Opuscula Sancto Patricio Adscripta.
Following the restoration of Charles II, Ware returned to Dublin where he was re-elected as M.P. for Dublin University. He remained on close terms with Ormond, who frequently visited him for consultations at his home in Castle Street.
1664 saw the publication of Venerabilis Bedae Epistolae Duae and Rerum Hibernicarum Annales ab Anno Domini 1485 ad Annum 1558. In the following year, which saw the publication of De Praesulibus Hiberniae Commentarius, he began a brief though fruitful collaboration with Dubhaltach MacFhirbhisigh. It was later stated that "He always kept in his House an Irish Amanuensis to interpret and translate the Language for him, and at the Time of his Death one Dubley Firbisse served him in that Office.
Ware's children include James (died 1689), Robert (died 1696) and Mary (died 1651). Mary married Sir Edward Crofton in 1647, he being a nephew of Thomas Crofton of Longford, Tireagh, Co. Sligo (another Thomas Crofton, of this family, killed Mac Fhirbhisigh in January 1671). A first cousin of Thomas Crofton of Longford was Catherine Crofton, daughter of John Crofton of Lisdorne, Co. Roscommon; Catherine was married to Reverend Joseph Ware, Dean of Elphin, who is believed to have been a younger brother of Sir James. James Ware junior had one daughter Mary (1651-1722) who married firstly Alexander Fraser and secondly Sir John St Leger, Baron of the Court of Exchequer (Ireland): she was described as a lady of great wealth and "questionable virtue".
Ware died on Saturday 1 December 1666, aged seventy-two years and five days. He was buried in St Werburgh's Church, Dublin.
His works were republished by his son Robert and by the husband of his great-granddaughter Mary – Walter Harris, the first serious historian of Dublin.