The Creed promulgated by the Nicene Council did not settle the issue of Arianism; indeed, its definition of the relationship of the Son to the Father served only to fan the fires of controversy which were to burn fiercely for the next fifty years. It appears that Eustathius of Antioch, one of the signatories of the Letter of the Synod of Antioch which had provisionally excommunicated Eusebius of Caesarea, was not convinced of the latter's honesty and sincerity in subscribing his name, after a day's delay, to the Nicene definition, and continued to charge him with holding Arian views. Eusebius counter-attacked by charging Eustathius with Sabellianism. At a Synod held in Antioch, shortly after the Arians regained favour with the Emperor, Eustathius was deposed and exiled from Antioch. The deposition, on whatever grounds it was achieved, marked the beginning of the Arian policy of forcing a reversal of the decision of Nicaea by removing, one by one, the supporters of the Nicene definition.

Having removed Eustathius, the Arians turned their attention to Athanasius who had succeeded to the see of Alexandria a few months after the Council had ended, and in A.D. 335 Athanasius was exiled to Treveri in Gaul, after being found guilty by an Arian Synod held at Tyre of murdering Arsenius, a Melitian bishop, and of violently treating Ischyras, a Melitian presbyter, while the latter was in the very act of celebrating the Eucharist. It was possibly during this period of exile that Athanasius composed his three Orations against the Arians, which are the main source for the study of Athanasius' use of St John's Gospel in the refutation of the teachings of the Arians.