Vincenzo Natali wanted to make the film after the success of his first film Cube (1997). But the technology hadn't reached the point yet where Natali could give it the treatment it deserved. He shelved the project for ten years, after which he felt the time had finally come.
The main scientist characters, Clive and Elsa, are named as an homage to Bride of Frankenstein (1935), a similarly themed movie. Colin Clive played Dr. Henry Frankenstein while Elsa Lanchester played Mary Wollstonecraft Shelley/The Bride.
The gestation cell that Clive and Elsa use to splice Dren has the acronym BETI placed conspicuously on the front of the machine. BETI stands for Biomechanical Extroutero Thermal Incubator. However, interestingly the word "BETI" means daughter in Urdu and Hindi, spoken in large parts of Pakistan and India.