Allah Nazar said he was still leading the insurgency. A Baloch Liberation Front (BLF) spokesman said the video was shot this month.
Government officials told the BBC they were working to verify the video.
The rebels say they will oppose planned Chinese investment in the region.
"For quite some time the government has been issuing propaganda about my death. This is a blatant lie," said Nazar, who vowed to continue to fight.
In September, Balochistan Home Minister Sarfaraz Bugti said the government believed Nazar had been killed in a major military operation in Awaran district in August.
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The BLF is one of several separatist groups that have been fighting against government forces in the mineral-rich province for almost a decade.
Despite its natural resources, the province is still the least developed part of the country. Separatists say it is being exploited by other parts of Pakistan.
The rebels have carried out kidnappings and bombings against government and security targets, including gas pipelines and other infrastructure.
Human rights groups accuse the military of a campaign of kidnapping, torture and extrajudicial killings of suspected separatists.
"They've turned Balochistan into a slaughterhouse," Nazar said.
He also said his forces would block China's planned $46 billion economic corridor, which passes through the country and ends in the province's Gwadar port.
Nazar said the project was causing Baloch people to be evicted from their homes.
The rebel leader, a doctor from the provincial capital of Quetta, is the most prominent Baloch separatist leader still fighting in the country.
Others are in exile, including Baramdagh Bugti, who in August said he was ready to negotiate with the government.
A separatist leader from Pakistan's southwestern Balochistan province, who the government believed it had killed in August, has appeared in a new video.