Quesnel joined the French Oratory (a religious society of secular priests) in 1657 and was ordained in 1659. His Jansenist sympathies led to his banishment from Paris in 1681, and three years later he was expelled from the Oratory for refusing to accept the anti-Jansenist decrees it promulgated. He fled to Brussels where he lived with the exiled Antoine Arnauld, champion of the Jansenist resistance, until Arnauld's death in 1694. In 1703 he was arrested but soon escaped to Amsterdam, where he finally settled.

Quesnel's Nouveau Testament en français avec des réflexions morales (1692; “New Testament in French with Thoughts on Morality”) was a major contribution to the literature of Jansenism, but it caused serious repercussions. It rekindled doctrinal conflicts between the Jansenists and the papacy, which were further complicated by the intervention of Louis XIV. Pope Clement XI's bull Unigenitus (1713)—prompted by Louis—condemned 101 sentences from the Réflexions morales, yet Quesnel never admitted that his opinions were heretical. Quesnel's correspondence was edited by A. Le Roy (2 vol., 1900).