Zoo officials say 24 monkeys, one bird and a zoo employee were inside at the time.
There was significant damage to the roof and side of the Exotic Animal House, which houses mostly primates.
Zoo employees scrambled to rescue and capture the animals after the impact. They were taken out one by one in animal carriers and transported to another building at the zoo.
The animals will be temporarily relocated to other zoos in the region.
Fortunately there were no injuries to report, but three small primates are being watched closely.
"We have a gentleman who actually sits at this building," said William Montgomery of the Delaware Zoological Society. "He luckily is okay. No animals injured, I'm sure they're stressed out beyond belief, but no obvious injuries from the tree."
It's unclear if Friday's rainy weather caused the tree to fall. People in the area at the time said there was no significant wind. The tree shows evidence of significant rotting in the trunk.
Officials say a crane will be brought in to hoist the tree off the building. The building will remain closed for some time.
The zoo's website says the zoo itself will remain closed until further notice.
The zoo is located at 1001 North Park Drive in Wilmington.