The Vienna-based security body said it had not heard from its team in the industrial region of Lugansk since yesterday evening when it was stopped "by armed men" at a roadblock in the town of Severodonetsk.
"The team was comprised of four international monitors and a Ukrainian translator," the OSCE's Special Monitoring Mission for Ukraine said in a statement posted on Facebook.
"The team was travelling in two cars."
The organisation added that another four members who were detained by separatist rebels in the neighbouring Donetsk region on Monday were still missing.
Another pro-Russian rebel from the little-known Southeastern Front claimed in a statement issued to the Interfax news agency that his men had detained the team in Lugansk.
But a spokesman for the self-proclaimed "Lugansk People's Republic" contacted by AFP denied that the group was being held against its will.
"Nobody arrested the four OSCE observers," Volodymyr Inogorodskiy told AFP.
"They finished their work late yesterday night in Severodonetsk and we advised them not to leave the city because of the (security) situation," said the spokesman.
But their mission has been treated with suspicion by the rebels throughout their seven-week insurgency.
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