The 34-year-old midfielder stepped away from international duty after Euro 2020 when Die Mannschaft were knocked out by England in the quarter-finals.
However, Kroos announced at the end of last month that he is "in the mood" to come out of retirement and represent Euro 2024 host nation Germany this summer.
The 2014 World Cup winner, who has earned 106 international caps, has now officially been included in Nagelsmann's latest squad ahead of friendlies away against France on March 23 and at home to the Netherlands on March 26.
Meanwhile, Bayern Munich midfielder Aleksandar Pavlovic is one of six players along with Maximilian Beier, Deniz Undav, Waldemar Anton, Maximilian Mittelstadt and Jan-Niklas Beste to receive their first senior call-up to the Germany squad.
Munich-born Pavlovic is of Serbian decent and although the Serbian FA sent a delegation to meet the midfielder and his family earlier this month, the 21-year-old has opted to represent Germany.
Another notable inclusion in Nagelsmann's squad is Bayern goalkeeper Manuel Neuer, who was omitted from the previous Germany set-up at the end of last year to focus on club duties.
The 37-year-old is expected to regain the number-one jersey from Marc-Andre ter Stegen, while Bernd Leno and uncapped Oliver Baumann are two more shot-stoppers at Nagelsmann's disposal.
In defence Chris Fuhrich and Robin Koch have both returned to the senior squad, the latter receiving his first call-up for nearly three years.
Barcelona's Ilkay Gundogan will continue as Germany captain and is one of only three midfielders out of the eight called up, along with Kroos and Jamal Musiala, who has earned at least 25 senior caps.
Arsenal's Kai Havertz is considered as one of five attacking options to Nagelsmann despite being experimented in a hybrid left-back role last year, and is joined in the squad by Thomas Muller and Niclas Fullkrug.
Standout names who have not been selected in Nagelsmann's latest squad include Leroy Sane, Serge Gnabry, Timo Werner, Leon Goretzka, Emre Can, Julian Brandt, Mats Hummels, Niklas Sule and Nico Schlotterbeck.
Germany will travel to the Groupama Stadium in Lyon to face France a week next Saturday, before returning home to take on the Netherlands at Deutsche Bank Park in Frankfurt three days later.
Germany's 26-man squad in full:
Goalkeepers: Oliver Baumann (Hoffenheim), Bernd Leno (Fulham), Manuel Neuer (Bayern Munich), Marc-Andre ter Stegen (Barcelona)
Defenders: Waldemar Anton (Stuttgart), Jan-Niklas Beste (Heidenheim), Benjamin Henrichs (RB Leipzig), Joshua Kimmich (Bayern Munich), Robin Koch (Eintracht Frankfurt), Maximilian Mittelstadt (Stuttgart), David Raum (RB Leipzig), Antonio Rudiger (Real Madrid), Jonathan Tah (Bayer Leverkusen)
Midfielders: Robert Andrich (Bayer Leverkusen), Chris Fuhrich (Stuttgart), Pascal Gross (Brighton & Hove Albion), Ilkay Gundogan (Barcelona), Toni Kroos (Real Madrid), Jamal Musiala (Bayern Munich), Aleksandar Pavlovic (Bayern Munich), Florian Wirtz (Bayer Leverkusen)
Attackers: Maximilian Beier (Hoffenheim), Niclas Fullkrug (Borussia Dortmund), Kai Havertz (Arsenal), Thomas Muller (Bayern Munich), Deniz Undav (Stuttgart)