Analogues of vitamin D3 are extensively used in the treatment of various illnesses, such as osteoporosis, inflammatory skin diseases, and cancer. Functional testing of new vitamin D3 analogues and formulations for improved systemic and topical administration is supported by sensitive screening methods that allow a comparative evaluation of drug properties. As a new tool in functional screening of vitamin D3 analogues, we describe a genomically integratable sensor for sensitive drug detection. This system facilitates assessment of the pharmacokinetic and pharmadynamic properties of vitamin D3 analogues. The tri-cistronic genetic sensor encodes a drug-sensoring protein, a reporter protein expressed from an activated sensor-responsive promoter, and a resistance marker.