The so-called ban on "partial birth" abortion was a long-sought victory for the Christian right and abortion opponents in the run-up to the 2004 election.
But within minutes of the signing ceremony, a federal judge in Nebraska issued a temporary restraining order barring US Attorney John Ashcroft and the Justice Department from enforcing the new law against four doctors who practice in more than a dozen states.
A district judge in New York took a similar legal challenge under advisement after an almost three-hour hearing. Abortion rights groups have also asked a California court to block and ultimately overturn the new law.
Bush, an evangelical Christian, cast the ban in stark terms: as the government coming to the defence of the "innocent child" against "a terrible form of violence."
"Today, at last, the American people and our government have confronted the violence," Bush said.
Abortion rights groups countered that the ban would sacrifice women's health and rights for Bush's political gain, and promised a court fight that could drag on for months if not years.
Bush vowed his administration would "vigorously defend this law against any who would try to challenge it in the courts."
He took the unusual step of signing the ban into law at a building named after Ronald Reagan, the former Republican president who opposed abortion, instead of at the White House.
"The ban will chill the practice of medicine and endanger the health of countless women"
Democratic presidential candidate
Outside, dozens of protesters denounced him with signs that read: "Keep abortion legal" and "We trust our doctors more than we trust Bush." The American Civil Liberties Union called it "a deceptive and dangerous" measure.
In a statement, Democratic presidential candidate Howard Dean, a doctor, called it a "dark day for American women" and warned the ban would "chill the practice of medicine and endanger the health of countless women."
White House spokesman Scott McClellan said the ban was a "practical" step to reduce US abortions and Bush would support additional efforts to end abortions, including measures to promote adoption and sexual abstinence.
In drafting the law, the US Congress determined such an exception was not necessary for health reasons, but did include a more narrow one to save a woman's life.
Under the bill, a doctor could face up to two years in prison as well as civil lawsuits for performing a "partial birth" abortion, defined as intentionally killing a foetus that has been partially delivered.