Catalan trade unions and associates called for the action due to “the grave violation of rights and freedoms” seen when heavily armed Spanish officers clashed with voters on Sunday.
Madrid, which deemed the referendum illegal, deployed hundreds of officers in a last desperate attempt to stop the ballot from going ahead.
But more than 2.2m Catalans were able to vote, with 90 per cent favouring freedom from Spain.
Still the brutal police crackdown left 893 people injured, as officers armed in riot-gear stormed polling stations, fired rubber bullets and pulled women by their hair.
Reacting to the violence, today’s strike will see public transport, schools and clinics in Catalonia closed.
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Barcelona’s famous football team FC Barcelona, the city’s public universities and contemporary art museum are also expected to strike.
The stoppage expected to have massive economic impact as Catonia accounts for a fifth of Spain’s economy.
Catalan leader Carles Puigdemont said: “I am convinced that this strike will be widely followed.”
Mr Puigdemont said on Monday he had not had any contact from the Spanish government regarding negotiations and called on the European Union to mediate.
And he has not ruled out making a unilateral declaration of independence.
The Catalan leader also announced the creation of a “special commission” to investigate claims of abuse of basic rights, adding legal action would be taken “against those responsible for violence during Sunday’s referendum”.
Spain’s constitutional court has insisted any independence vote in Catalonia is illegal, while Prime Minister Mariano Rajoy refused to acknowledge the ballot on Sunday.
Speaking after the vote, Mr Rajoy said: “There has not been an independence referendum in Catalonia today.”
The European Commission criticised the police violence but said any independent Catalonia would find itself outside the bloc.
United Nations rights chief Zeid Ra’ad Al Husseind said he was “very disturbed” by the rest, while EU President Donald Tusk urged Madrid to avoid “further use of violence”.
European Commission spokesman Margaritis Schinas said: “We call on all relevant players to now move very swiftly from confrontation to dialogue. Violence can never be an instrument in politics.”