Koulibaly is considered one of Europe's best centre-backs at a time when the position is struggling for top-level players and Manchester United are among the clubs to have followed his progress closely.
The Senegalese defender changed agents earlier this season, often an indication that a player is preparing for a move, but Ancelotti was keen to nip exit talk in the bud ahead of Napoli's Europa League quarter-final second leg against Arsenal.
"Koulibaly is the best centre-back in the world - and he has more room for improvement," the former Chelsea boss said.
"He is worth €150 million and he is non-transferable.
"Napoli do not need to sell, the president does not want to sell and the question does not arise."
Napoli president Aurelio De Laurentiis has a reputation for getting good prices for his players, and previously insisted that stars like Gonzalo Higuain and Jorginho would not be allowed to leave before their eventual transfers.
The same could happen with Koulibaly, but the fear among the 27-year-old's entourage is that the club might price him out of a dream move and life-changing salary in the Premier League.