Calvert-Lewin has been in fine form for Leeds this season with 10 Premier League goals, while Mainoo has enjoyed a return to regular football with Manchester United following the appointment of Michael Carrick as manager.
The striker last appeared for England five years ago in the quarter-finals of the delayed Euro 2020 tournament - and Mainoo hasn't been selected since September 2024.
The duo is part of several surprise selections as manager Thomas Tuchel gets one last look at potential players ahead of the 2026 World Cup.
Brighton goalkeeper Jason Steele is involved with a view to becoming the Three Lions' training goalkeeper in the summer.
Also included in the squad are Everton's James Garner, Newcastle left-back Lewis Hall, Manchester United defender Harry Maguire, AC Milan star Fikayo Tomori and Tottenham striker Dominic Solanke.
Several staples of Tuchel's England set-up also return, such as Harry Kane, Bukayo Saka, Jude Bellingham and Jordan Pickford, among other big names.
Trent and Watkins overlooked
There is no place in the squad, however, for Ollie Watkins or Trent Alexander-Arnold, while Reece James misses out through injury.
England have named a larger squad, consisting of 35 players, for friendlies against Uruguay on March 27th and Japan four days later on March 31st.
"We decided to divide it into basically two camps, almost, so we bring players in that we haven’t seen who haven’t played so much to open up the picture, and the competition for plane tickets to the US," Tuchel explained.
"Then from Friday and Saturday, a group of players will come into camp - 10 or 11 players who get a rest before, and we will then go with a new group and mix of players into the match against Japan."
The Three Lions enjoyed an unbeaten World Cup qualifying campaign, winning every match without conceding a goal.
The March international break represents a chance for those on the fringes of the side to stake their claim ahead of the World Cup this summer.
Full 35-man England squad
Goalkeepers: Dean Henderson, Jordan Pickford, James Trafford, Aaron Ramsdale, Jason Steele
Defenders: Dan Burn, Marc Guehi, Lewis Hall, Ezri Konsa, Tino Livramento, Harry Maguire, Nico O'Reilly, Jarell Quansah, Djed Spence, John Stones, Fikayo Tomori
Midfielders: Elliot Anderson, Jude Bellingham, James Garner, Jordan Henderson, Kobbie Mainoo, Declan Rice, Morgan Rogers, Adam Wharton
Forwards: Jarrod Bowen, Dominic Calvert-Lewin, Eberechi Eze, Phil Foden, Anthony Gordon, Harry Kane, Noni Madueke, Cole Palmer, Marcus Rashford, Bukayo Saka, Dominic Solanke