The two boys were shot dead when they were mistaken for insurgents during an operation in the northwest of Uruzgan on February 28, ISAF commander, US general Joseph Dunford, said in a statement.
"I offer my personal apology and condolences to the family of the boys who were killed," Dunford said in the statement. "The boys were killed when Coalition forces fired at what they thought were insurgent forces," he said.
A team of Afghan and ISAF investigators visited the village on Saturday and met with local leaders, Dunford said.
Australian forces, who are deployed in Uruzgan, said earlier there had been an "operational incident" in the province but gave no details.
On February 13 a NATO air strike requested by Afghan forces killed 10 people - including five children and four women - in the eastern province of Kunar, prompting Karzai to ban his troops from requesting foreign air strikes.
Two weeks later he halted all special forces operations in the central province of Wardak after a series of allegations involving U.S. special forces soldiers and Afghan men said to be working with them.