Operating legally within the borders of the Second Polish Republic since 1924, it continued its functions without interruption even after Vilnius became part of the Soviet Union.

The monastery occupies a complex of one-story buildings in a courtyard behind an ornate entrance gate at 10 Aušros Vartų Street [pl].

In 1597, the brotherhood succeeded in obtaining a royal privilege to build a new Orthodox church in Vilnius on land owned by sisters Theodora and Anna Wołowicz.

There, they established a male monastery, with Longin Karpowicz, a printer in the brotherhood's workshop who took the monastic name Leoncjusz [pl], becoming its first superior.

[7] The monastery sustained itself through the monks' work in the orchard and farm and with support from lay donors, mainly from the noble ranks of the Brotherhood of the Holy Spirit, such as Chamberlain Bogdan Ogiński.

On the left side of the courtyard was a one-story building housing the living quarters for the superior and deputy of the monastery and the office.

The Brotherhood of the Holy Spirit[9] and the school it organized, known for its high standards and intended as an alternative to Jesuit colleges, operated in adjacent buildings.

[11] It was one of two monasteries in the Polish–Lithuanian Commonwealth managed by the diocesan bishop of the respective diocese (in this case, Vilnius) or an archimandrite appointed by the Metropolitan of Kyiv in agreement with the Brotherhood of the Holy Spirit.

[12] In 1629 and 1637, the monastery was attacked by groups of Union supporters aiming to eliminate the last strong center of disunity on Lithuanian lands.

[4] The monks also faced difficulties obtaining permission to build a new brick church, which they eventually received through a privilege granted by King Władysław IV Vasa in 1632.

[8] In the second half of the century, during the wars of the Polish–Lithuanian Commonwealth with Sweden and Russia, the monastery suffered multiple times from military actions or was looted.

[4] King John II Casimir Vasa had no doubts about the community's loyalty and confirmed their faithfulness to the Commonwealth in a letter to the city authorities, instructing them to exempt the monastery from military levies due to war losses.

[6] The city authorities agreed to the monastery's reconstruction only in 1751, with the condition that the new buildings would not stand out too much among the neighboring structures and that the gate facing the street would not be too richly decorated.

[8] In May 1797, Tsar Paul I recognized the Vilnius monastic community as a second-class monastery, which meant an annual government grant of 2,500 rubles.

[8] However, Paul's successor, Alexander I, considered closing the monastery in 1806 due to the lack of active Orthodox churches in Vilnius.

The Most Holy Synod strongly advised against closing the monastery, given its revered status and long tradition of strengthening Orthodoxy in Lithuania.

[8] His successor, Metropolitan Nikander Molchanov [pl], oversaw a general renovation of the entire complex,[8] directed by eparchial architect A.

[16] In the first year of World War I, the monks engaged in charitable activities, working in hospitals, and the church held services for Russian military victories.

The monastery buildings were undamaged during the war,[8] but the monks did not return until 1919, when a new Bishop of Vilnius, Eleutherius Bogoyavlensky, appointed by the Patriarch of Moscow, arrived.

[8] During the Second Polish Republic, the campaign to reclaim Orthodox church property initially did not threaten the monastery’s existence, as it had never been affiliated with Catholic institutions.

[8] In 1924, a request from the new Metropolitan of Warsaw, Dionysius Waledyński, for official recognition of the monastery received a favorable response from the Ministry of Religious Affairs and Public Education.

[16] Lacking arable land, the monastery could not sustain itself through farming but relied on rental income and voluntary donations from the faithful for maintenance and regular building upkeep, such as updating electrical installations in 1928 and purchasing new candleholders for the church.

[19] In 1929, Roman Catholic Archbishop of Vilnius, Romuald Jałbrzykowski, filed a lawsuit for the recovery of both the Holy Spirit Church and the monastery.

[8] Under Nazi occupation following Germany's attack on the Soviet Union, the monastery buildings hosted three-semester courses for future priests.

After 1982, Archbishop Wiktoryn (Bielajew) initiated the construction of an elevator leading to the church towers and renovated the monks' living quarters.

The buildings also house the office of the Vilnius and Lithuanian diocese, as well as its secretariat responsible for the charitable activities conducted by Orthodox priests.

In July of that year, during the feast of the Descent of the Holy Spirit, the 400th anniversary of the continuous operation of the monastery church was celebrated with the participation of Patriarch Alexy II of Moscow.

[25] The monastery buildings, inhabited by monks and occupied by the diocesan office and library, have been preserved to this day in the same form as they were rebuilt after the fire in the 18th century.