Zoo officials conducted an ultrasound and found no evidence of a fetus, the Washington Times reported Saturday.
The disappointing news means Tai Shan, who turns two Monday, will remain the National Zoo's only panda cub for the foreseeable future.
Tai Shan is the zoo's biggest attraction and more than 2 million visitors have watched him grow from just a few ounces to 125 pounds.
Zoo officials hoped for another cub after Mei Xiang was artificially inseminated in April with semen taken from Gao Gao, a giant panda at the San Diego Zoo.
2014: The Year in Music [PHOTOS]