Breast cancer is one of the biggest killers of women in the UK, claiming 10,000 lives a year, despite recent advances in treatment and detection.
The statistical study from the Institute of Cancer Epidemiology is the most compelling evidence yet of a link between the two.
It still leaves doctors with little idea as to why there might be a connection, although there is speculation that altered exposure to light during the hours of darkness may trigger hormonal changes which increase the risk.
Breast cancer is one of the biggest killers of women in the UK, claiming 10,000 lives a year, despite recent advances in treatment and detection.
The Danish study looked at more than 7,000 women between the ages of 30 and 54 years.
Full employment histories were reconstructed dating back to 1964, and the results were altered to take account of other risk factors such as alcohol consumption, and age at the birth of first and last children.
The researchers found that women who had worked predominantly at night for at least six months in their working life had an increased incidence of breast cancer.
Statistically, they were 50% more likely to develop the disease.
Biggest study yet
All the statistical evidence was less strong, there was a trend towards longer periods spent working at night increasing that risk yet further.
The study is the largest ever to look at this particular risk factor - and the most statistically sound.
Dr Tim Key, a senior epidemiologist at the Imperial Cancer Research Fund, said that although the suspicion that night-working was a factor had been around for a while, little evidence existed.
He said: "This hypothesis (that disruption of hormone secretion patterns by disturbed exposure to light or to electromagnetic fields might increase breast cancer risk) was put forward in the 1980s.
He added: "The findings of this new study could be interpreted as supporting the possibility that some lifestyles might increase breast cancer risk, perhaps by altered exposure to light and altered sleep patterns, which might increase risk partly through reducing levels of the hormone melatonin.
"However, this is currently only speculative, there could be other explanations for the findings, and more research is needed to improve understanding of this topic."
The findings were contained in the journal Epidemiology.