The Chalk River project centred on the NRX reactor, which provided the fission process needed to convert Uranium-238 into Plutonium-239 in the fuel rods. By spring 1950, the extraction plant was pulling 50 grams of plutonium per week from thorium-loaded tubes surrounding the fuel rods. The work was dangerous. On December 13, 1950, an explosion in stored ammonium nitrate concentrate killed operator Stephen Whalen instantly and hospitalized four others. In 1952, the Canadian government created Atomic Energy of Canada Limited (AECL) to steer the facility toward peaceful applications. AECL took over operations from the National Research Council and began producing nuclear material for medical and scientific use. Despite the shift in mandate, plutonium extraction quietly continued. From 1955 to 1985, Chalk River supplied spent reactor fuel containing plutonium to the U.S. Department of Energy for nuclear weapons production.

On December 12, 1952, the NRX reactor experienced a power excursion and partial loss of coolant. Mechanical failures compounded human errors: control rods could not be lowered into the core, and three rods that did not reach their destination were pulled back out by accident. The fuel rods overheated into meltdown. Hydrogen explosions ripped through the reactor building, blowing the seal of the reactor vessel four feet into the air. Radioactive water flooded the cellar and was eventually dumped in ditches near the Ottawa River. Among the 26-person cleanup team was a young U.S. Navy officer stationed in Schenectady, New York: Jimmy Carter, the future President of the United States. Fourteen months later, the repaired NRX was running again. A second accident struck in 1958 when a fuel rod ruptured in the newer NRU reactor. A robotic crane extracted a rod of metallic uranium, which caught fire and broke, the largest piece falling back into the containment vessel while still burning. Scientists and maintenance workers extinguished the blaze by sprinting past the smoking opening in protective suits, hurling buckets of wet sand as they passed.

For decades, Chalk River's reactors produced something the world depended on far more than weapons-grade material: medical radioisotopes. At its peak, the facility generated roughly one-third of the global supply and half the North American supply of isotopes used in diagnostic imaging and cancer treatment. This role made the facility's operational status a matter of international concern. In 2007, the Canadian Nuclear Safety Commission ordered the NRU reactor shut down over safety upgrades, prompting Parliament to pass emergency legislation overriding the decision and authorizing a restart within days. Prime Minister Stephen Harper publicly criticized the regulator, arguing the shutdown "jeopardized the health and safety of tens of thousands of Canadians" who depended on the isotopes. When the NRU went offline again in 2009 due to a heavy water leak, only one of the world's four other regular isotope-producing reactors was operational, triggering a worldwide shortage.

Chalk River's contributions extended well beyond isotope production. Bertram Brockhouse conducted his pioneering work in neutron spectroscopy at CRL from 1950 to 1962, research that earned him the 1994 Nobel Prize in Physics while he was a professor at McMaster University. Sir John Cockcroft, himself a Nobel laureate, served as an early director of the facility. Canada's first nuclear power plant went online in 1962 near the Chalk River site: the Nuclear Power Demonstration reactor, a proof of concept for the CANDU reactor design that would become one of the world's most successful nuclear technologies. The CANDU's use of natural uranium fuel and heavy water moderation made it distinctive among reactor designs and established Canada as a significant player in civilian nuclear power.

The NRU reactor was shut down for the last time at 7 p.m. on March 31, 2018, entering a state of storage ahead of decommissioning. But Chalk River is far from finished. In 2016, the Canadian government allocated 1.2 billion dollars over ten years to decommission 120 aging buildings and construct modern replacements, completed starting in 2020 as the Canadian Nuclear Laboratories Research Facilities. The ZED-2 zero-energy research reactor, operational since 1960, continues to run. In May 2023, it was announced that the world's first micro-modular reactor, developed by Global First Power, will be built at Chalk River to power the campus as a demonstration unit. The plan envisions shipping-container-sized microreactors eventually being transported to remote northern communities to replace diesel generators, saving an estimated 200 million litres of fuel. Eight decades after its founding as an atomic bomb factory hidden in the Ontario woods, Chalk River is reinventing itself once more.