In 1558, Antonio was introduced into the service of Duke Cosimo I de' Medici by the secretary Iacopo Guidi, his relative and fellow native of Volterra, who would later become the bishop of Atri and Penne.

[2] During this period, Serguidi carried out delicate diplomatic missions: in 1569, he was the extraordinary envoy of Cardinal Ferdinando to thank the pope for granting the grand ducal title.

He returned to Rome several times: in 1572 to secure benefits for the Order of St. Stephen, in 1574 to negotiate over the Orsini’s claims on Pitigliano, in 1575 to address the reception of Tuscan ambassadors in the Sala Regia, and in 1581 to congratulate the convalescing Pope Gregory XIII.

Serguidi was to inform Philip II that Grand Duke Francesco had shown a strong inclination to side with Spain and hoped to “deserve that His Majesty would finally declare his intention in this matter of the title and show that he warmly sheltered us under his wings”.

[3] In 1577, Serguidi once again journeyed to Spain, this time to pave the way for the visit of Pietro de' Medici, Francesco's brother and Cosimo's youngest son, to Philip II's court.

Initially, Serguidi communicated with Antonio Pérez, Philip II's influential secretary, regarding matters of the Grand Duchy's foreign policy.

Subsequently, he directly addressed Philip II, affirming that "all our faith – as Francesco wrote – is placed in him, and we will forever navigate by his north star."

Initially, Bartolomeo Concini held a prominent role, but after 1576, possibly due to his advanced age, the handling of important matters shifted to his son-in-law, Serguidi.

In November 1587, Ferdinando established a three-member secretariat, including Serguidi alongside Belisario Vinta and Pietro Usimbardi (who assumed the role of first secretary).