Jefferson had been fighting the case in court for the last 16 months.
A three-judge panel of the 4th U.S. Circuit Court of Appeals rejected Jefferson's claims that a federal grand jury received evidence that violated his constitutional right to legislative immunity.
The 2007 indictment alleges that Jefferson received more than $500,000 in bribes and demanded millions more between 2000 and 2005, including $90,000 he received from an FBI informant that was later found in the freezer of his Washington home.
Jefferson could further delay a trial by appealing to the U.S. Supreme Court.
If convicted of all charges, he faces up to 235 years in prison.
Jefferson has pleaded not guilty.