In calling for the protest, al-Sadr said in a statement: "In order to end the occupation, you will go out and demonstrate."
Thousands of Shia responded on Sunday by riding buses or cars into Najaf.
"The chances of success [in Iraq] are essentially zero because the Iraqi people have no voice"
Non Sequitur, Cadiz, Spain
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The Baghdad-Najaf road was packed with hundreds of vehicles crammed with passengers waving Iraqi flags and chanting religious and anti-US slogans.
"No, no, no to America ... Muqtada, yes, yes, yes!" they chanted.
Witnesses in some southern towns said Iraqi police were trying to stop al-Sadr's supporters from getting to Najaf.
Also on Sunday, four American soldiers were killed in attacks south of Baghdad while another two died from wounds suffered in operations north of the capital, the US military said.
The toll makes it a deadly weekend for American troops after four soldiers were killed in an explosion near their vehicle in Diyala province north of Baghdad on Saturday.
Al-Sadr urged the al-Mahdi Army militia and Iraqi security forces to stop fighting in the city of Diwaniya, in southern Iraq, and to stop playing into the hands of the US forces.
Iraqi and US forces have clashed with militiamen in Diwaniya since launching an operation on Friday to take control of the southern city.
The Pentagon says the militia is the greatest threat to peace in the country.
On Sunday, a series of attacks in and around Baghdad killed more than 20 people and wounded dozens.
In the deadliest attack, a car bomb killed 17 people and wounded 25 in the town of Mahmudiya, 30km south of Baghdad, city officials said.
Another seven people were killed in a suicide car bomb in Baghdad.
And in the district of Adhamiya, a policeman was killed and two were wounded when a roadside bomb exploded near their patrol.