Sanders declined when asked on CNN's "Erin Burnett OutFront" to say whether or not his wife was under investigation by the FBI.
"My wife is about the most honest person I know," Sanders said.
Sanders' former campaign manager, Jeff Weaver, confirmed Jane Sanders has hired lawyer Larry Robbins to represent her in the possible probe. Weaver said the FBI had not contacted either Jane or Bernie Sanders in connection to any such investigation. A person close to the senator said Sanders was represented by Rich Cassidy, a longtime lawyer for the family.
Both the FBI field office in Albany, New York and the US Attorney's Office in Vermont would not comment, and the Justice Department has not yet responded to a request for comment on the matter.
The senator's wife was the president of Burlington College from 2004-2011. The college announced its closure
a year ago due to debt it had taken on from a real estate deal the school made during her tenure. CNN reviewed the loan application for the deal, which was signed by Jane Sanders. The loan application stated the college would take in millions of dollars worth of pledged donations, many of which never materialized.
Vermont lawyer Brady Toensing, a former state Republican vice chair who was once Trump's campaign chair in the state, has leveled accusations of bank fraud against the couple and alleged the senator may have pressured a Vermont bank to speed up the loan process. Toensing made these accusations five years after Jane Sanders left the college.
In the interview on Tuesday, Sanders claimed Toensing instigated the potential probe for political reasons, saying it began five years after his wife's time at the college but just as his presidential campaign was going on -- "coincidentally, no doubt," the senator said.
"I think it's fairly pathetic that when people are involved in public life, it's not only that they get attacked, but it's their wives and their families that get attacked," Sanders said.
Toensing told CNN on Sunday that his hope "is for a fair, full and impartial investigation."
Sanders also defended his wife's stewardship of the university.
"When she came to that college, it was failing financially and academically," Sanders said. "When she left it, it was in better shape than it had ever been."
Sanders declined to answer further questions about the matter, other than to say he would "let it play out" and again said an attack on a politician's wife or family was "pathetic."
Yves Bradley, the chair of the Burlington College Board of Trustees, told CNN he was informed of an FBI investigation by the college's dean of operations over a year ago and that he believed the investigation's focus to be the the college's disastrous land deal.
Bradley said Burlington College had been subpoenaed as part of that investigation.
Dr. Ron Leavitt, a former board of trustees member, told CNN that FBI agents had interviewed him in March and that school records had exaggerated a $30,000 donation he pledged to the school in 2010.
Leavitt said the agents never asked him about Jane Sanders.