Mr Trump also insisted he and his campaign had not colluded with Moscow in last year's election, and shifted blame on the Justice Department and his Democratic rival Hillary Clinton.
"I never asked Comey to stop investigating Flynn. Just more Fake News covering another Comey lie!" Mr Trump said in a tweet.
But his position was complicated by another Twitter post in which he indicated he had fired Flynn because the national security chief had been untruthful not just to Vice President Mike Pence but to the FBI as well.
That comment appeared to indicate Mr Trump was acknowledging he knew at the time of Flynn's firing in February that he had lied to the bureau's agents.
"If that is true, Mr. President, why did you wait so long to fire Flynn?" asked Representative Adam Schiff of California, the top Democrat on the House Intelligence Committee.
"Why did you fail to act until his lies were publicly exposed? And why did you pressure Director Comey to 'let this go?'"
White House officials, however, told The New York Times that Trump was only referencing Mr Flynn's guilty plea for lying to the FBI about his conversations with then Russian ambassador Sergey Kislyak over sanctions president Barack Obama slapped on Russia for election meddling.
And two people briefed on the matter said the Twitter post was in fact written by Mr Trump's personal lawyer John Dowd, who apologised to the White House for the tactless language.
What the president knew, and when
After he was fired himself in May, Mr Comey testified under oath before a Senate panel that, a day after Mr Flynn's firing, Mr Trump asked him to drop an investigation into the former national security advisor.
A lingering part of the drama has been that after the White House learned through the Justice Department that Flynn lied to the White House about discussing sanctions with the Russian ambassador, Mr Trump still waited 18 days to fire him.
Mr Trump said he had the Russia probe in mind when he fired Mr Comey. The move backfired and led the Justice Department to appoint Robert Mueller as special counsel.
Mr Mueller's focus goes beyond possible collusion with Russia to business dealings and whether Mr Trump himself tried to thwart the investigation.
US media reported that senior FBI counterintelligence official Peter Strzok was removed from the investigation over the summer for sending text messages critical of Mr Trump.
Mr Trump also retweeted another damaging Sperry post critical of Mr Strzok's boss, FBI Deputy Director Andrew McCabe.
Earlier, Mr Trump renewed focus on the Justice Department's handling of the Clinton probe.
"So General Flynn lies to the FBI and his life is destroyed, while Crooked Hillary Clinton, on that now famous FBI holiday 'interrogation' with no swearing in and no recording, lies many times...and nothing happens to her? Rigged system, or just a double standard?" he wrote.
"Many people in our Country are asking what the 'Justice' Department is going to do about the fact that totally Crooked Hillary, AFTER receiving a subpoena from the United States Congress, deleted and 'acid washed' 33,000 Emails? No justice!"
As he left for a day trip to New York on Saturday, Mr Trump again insisted his team had not plotted with Moscow to sway the election in his favor over Clinton, who won the popular vote but lost the all-important electoral college count.
"What has been shown is no collusion. There's been absolutely no collusion. So we're very happy," Mr Trump said.
Mr Comey himself seemed to be addressing the latest developments in an Instagram message: "To paraphrase the Buddha -- Three things cannot be long hidden: the sun; the moon; and the truth."
Tax win overshadowed
The explosive new developments in the Russia probe have overshadowed a major legislative win for Mr Trump: the Senate's passage of the most significant US tax overhaul in 31 years.
Both the Senate and House versions lower the corporate tax rate to 20 percent from 35 percent, and include more modest tax cuts aimed at individuals across all income levels.
Democrats argue that the plan is too expensive and will accommodate only the rich, and that it could ultimately impact cherished US entitlement programs like Medicare.
The Senate bill was, just 24 hours earlier, on the brink of collapse when a handful of Republican deficit hawks balked at the plan's $1.5 trillion price tag for 10 years.
Mr Trump hopes to sign a final bill before Christmas. That would be a much-needed victory for the president, who has delivered on hardly any of his major campaign promises, including repealing the health care law known as Obamacare.