"[Militao] underwent tests this Sunday... which revealed a minor muscle injury in his left thigh that will prevent him from playing," the CBF said in a statement.
Eder Militao, who missed September's fixtures due to a leg injury, first felt some discomfort against Atletico Madrid on September 29.
Brazil sit fifth in South America's qualification table after eight matches, with 10 points.
He is the second Madrid player to pull out of the Brazil squad, with Vinicius Junior also unfit to play after suffering a neck injury.
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