Masked gunmen wearing black clothing gunned down the protesters in Karbala, a Shia holy city in central Iraq, in one of the deadliest attacks since anti-government protests began this month.
More than 800 people are thought to be have been injured in the attack.
The early morning gunfire came as protesters took to the streets for the fifth day in a row to demonstrate against alleged government corruption, lack of services and economic hardship.
A total of 90 people have been killed since Friday, when the protests began, with a further 149 deaths earlier in the month during a separate wave of protests in the country.
It was not immediately clear who was behind Tuesday's attack, and protesters said they did not know whether the masked men were riot police, special forces or Iran-linked militias active in the country.
Security officials said the attack happened in Karbala's Education Square, just over a mile away from the area where the protesters had set up tents for a sit-in.