Fluorescent contrast agents can help identify the locations of tumors during surgery, but the contrast between tumors and healthy tissues can be reduced if a dye doesn’t specifically target a tumor. Researchers led by Matthew Bogyo of Stanford University report that they have developed new agents that boost fluorescence contrast in tumors by turning on only when two tumor-specific protein-cleaving enzymes are present (Nat. Biomed. Eng. 2020, DOI: 10.1038/s41551-020-00616-6). The so-called AND-gate contrast agents consist of a peptide scaffold with a fluorescent dye attached to a central portion and two quencher molecules attached to two arms. Each arm contains a cleavage site for a different tumor-specific enzyme. Fluorescence is activated only if both quenchers are removed, which requires both enzymes to be present and lessens the likelihood of observing a signal in healthy tissue. The researchers altered the selectivity of the contrast agents by incorporating different enzyme cleavage sites in the scaffold. They used the imaging contrast agent for fluorescence-guided robotic surgery to remove mammary tumors and metastatic lung tumors in mice. After excision, the contrast agent revealed residual cancer cells that couldn’t be seen with conventional white-light imaging.