Barcelona will be unable to sign players until January 2016 once the current transfer window shuts, after world governing body Fifa rejected the club's appeal against a transfer ban.
The Spaniards were given the 14-month ban after being found to have breached rules on signing international players under the age of 18.
It was suspended after they appealed, allowing them to sign Luis Suarez.
Barcelona will now take the case to the Court of Arbitration for Sport (Cas).
The ban will start in the next transfer window, early next year, which means their summer acquisitions will be unaffected.
And the Catalan club could, if they wanted, sign more players before the current transfer window ends on 31 August.
In a statement, the club said it would "continue defending its interests before the highest sporting authority, in this case the Court of Arbitration for Sport".
Barcelona added that the punishment was an "affront to the spirit" of their "world renowned" academy La Masia, which is regarded as being the foundation of the club's success in recent years.
Players are instilled with a philosophy of how the game should be played and many of the first team, such as Lionel Messi, Andres Iniesta and Xavi Hernandez, have graduated from the academy which is situated near the Nou Camp.
|Barcelona's summer signings|
|Luis Suarez (striker) - £75m from Liverpool||Marc-André ter Stegen (goalkeeper) - undisclosed from Borussia Monchengladbach|
|Thomas Vermaelen (defender) - £15m from Arsenal||Jeremy Mathieu (defender) - £15.8m from Valencia|
|Claudio Bravo (goalkeeper) - £7.2m from Real Sociedad||Jordi Masip (goalkeeper) - free from Barcelona B|
|Ivan Rakitic (midfielder) - undisclosed from Sevilla||Rafinha (midfielder) - recalled from loan at Celta Vigo|
The Spaniards have also been ordered to pay a fine of 450,000 Swiss Francs (£305,000) and been told by Fifa "to regularise" the situation of all minor players concerned.
"FC Barcelona is to serve a transfer ban which will see the club prevented from registering any players at both national and international level for two complete and consecutive transfer periods," read a Fifa statement.
Meanwhile, the Spanish Football Federation (RFEF) has been told to pay a fine of 500,000 Swiss Francs (£340,000) and told to "regularise its regulatory framework and existing system concerning the international transfer of minors in football" within a year.
The Catalans have signed eight players during the current transfer window for a total of about £130m.
They paid a reported £75m for former Liverpool striker Suarez, although Barcelona subsequently said the figure was nearer £65m, while defender Thomas Vermaelen was bought from Arsenal for £15m.
Barcelona were originally sanctioned by Fifa's disciplinary committee on 2 April for breaking the rules in the case of 10 minors.
After the club followed through on their promise to fight the ban, Fifa decided that because of the complexity of the case and the possibility of a further appeal to the Cas, it was unlikely to be concluded before the opening of the current transfer window on 1 July, and therefore suspended the punishment.
Fifa rules state that international transfers are only permitted for players over the age of 18 - unless the player in question meets one of three qualifying criteria.
Under-18s can move to a club in a different country if their parents move there for non-footballing reasons, if they are from another nation within the European Union or European Economic Area and aged between 16 and 18, or if they live within 100km of the club.
A Fifa investigation - centred on several players aged under 18 who were registered and played for the club between 2009 and 2013 - found that Barcelona and the RFEF were guilty of a "serious" infringement of the rules in relation to 10 players.
In April, Fifa told BBC Sport the identity of these players was confidential.