Commonly used in flow cytometry, multiplexed optical probes can diagnose multiple types of cell surface marker, potentially leading to improved diagnosis accuracy in vivo. Herein, we demonstrate the targeting of two different
tumor markers in models of disseminated
ovarian cancer. Two
ovarian cancer cell lines (SKOV3 and SHIN3) were employed; both overexpress
D-galactose receptor (D-galR), but only SKOV3 overexpresses HER2/neu. Additionally, fusion
tumors composed of SKOV3 and SHIN3/RFP were evaluated. Both
galactosyl serum albumin-rhodamine green (
GSA-RhodG), which binds D-galR, and trastuzumab-Alexa680, which binds HER2/neu, were administered to
tumor-bearing mice for in vivo fluorescence imaging and in situ fluorescence microscopy. In vivo fluorescence imaging depicted 64 of 69 SKOV3
tumors (94.2%) based on their dual spectra corresponding to both RhodG and Alexa680, while all 71 SHIN3
tumors (100%) were detected based on their single spectrum corresponding only to RhodG. All 59 SHIN3 and 36 SKOV3
tumors were correctly diagnosed with in situ microscopy. Additionally, in the mixed
tumor model, all
tumors could be depicted using the RhodG spectrum, but only SKOV3 components also showed the Alexa680 spectrum. In conclusion, multitargeted multicolor optical imaging enabled specific in vivo diagnosis of
tumors expressing distinct patterns of receptors, leading to improved diagnostic accuracy.