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MOSCOW—Police sealed off a central Moscow square and skirmished with anti-Kremlin activists Tuesday evening, detaining at least three Russian opposition leaders and dozens of followers who were part of a defiant, surging crowd that included observers from the European Parliament.

At least 160 protesters were hauled onto police buses in Moscow and St. Petersburg, police said, during the latest in a year-old series of rallies demanding freedom of assembly. That cause has become a focal point of political opposition, and the government's response is closely watched as a gauge of its tolerance for dissent.

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