The ability to employ mvpCu to image intracellular Cu(I) within tumors can be used for diagnostic purposes since high Cu(I) levels are linked to increased cancer aggression. To this end, 4T1 breast cancer cells were implanted in the mammary fat pads of BALB/c mice and once the tumor volume reached 100 mm 3 , we administered mvpCu systemically (1.0 mg/kg, 100 µL) for PA imaging. Because the turned over product is spectrally distinct from mvpCu, we were able to perform ratiometric imaging by irradiating each compound at 670 and 765 nm, respectively. We observed a 3.25-fold turn-on response relative to initial background scans, indicating successful detection of intratumoral Cu(I) (Figure ). As mentioned above, because the TPA trigger used in this design is only able to detect the labile Cu pool (defined as Cu associated with glutathione), we also performed ICP-MS measurements to examine whether the total Cu content was also higher in the tumors. Although not statistically significant, our results appear to support this notion (breast tumors = 3.88 ± 0.12 ppm vs. healthy beast tissue = 3.19 ± 0.81 ppm) (Figure ). When the performance of PACu-1 was compared head-to-head to mvpCu, it failed to detect intratumoral Cu(I) (Figure ). This finding demonstrates the powerful tumor targeting ability of our multivalent approach.