Francis spoke at the end of a Palm Sunday service, the first since 2019 in which the public had been allowed back in the square following two years of scaled-back services because of COVID-19 restrictions.
Some in the crowd put small Ukrainian flags at the tip of olive branches and a woman who read one of the prayers near the altar was dressed in the flag's blue and yellow colours.
"Put the weapons down! Let An Easter truce start. But not to rearm and resume combat but a truce to reach peace through real negotiations open to some sacrifices for the good of the people," Francis said.
Francis earlier evoked the horrors of war in his homily, speaking of "mothers who mourn the unjust death of husbands and sons...refugees who flee from bombs with children in their arms...young people deprived of a future....soldiers sent to kill their brothers and sisters."
Since the war began, Francis has only mentioned Russia specifically in prayers, such as during a global event for peace on March 25. But he has referred to Russia by using terms such as invasion and aggression.
Moscow dubs the action it launched on Feb. 24 a "special military operation" designed not to occupy territory but to demilitarise and "denazify" its neighbour. Francis has already rejected that terminology, calling it a war.
Palm Sunday commemorates the day the Gospel says Jesus rode into Jerusalem on a donkey and was hailed by the people, only to be crucified five days later.
It marks the start of Holy Week leading up to Easter Sunday in the Roman Catholic Church on April 17 this year. Ukraine is predominantly Orthodox. Orthodox Christians celebrate Easter a week later, on April 24.
A flare-up of pain in his knee forced Francis, 85, to skip the traditional procession from the obelisk at the centre of the square to the altar on the steps of St. Peter's Basilica.
He watched instead while seated at the altar, to where he was driven in a small car. He later limped as he said the Mass.
(Reporting by Ayman)