Optical fluorescence imaging is increasingly used to monitor biological functions of specific targets in small animals (1-3). However, the intrinsic fluorescence of biomolecules poses a problem when fluorophores that absorb visible light (350–650 nm) are used. Near-infrared (NIR) fluorescence (650–900 nm) detection avoids the natural background fluorescence interference of biomolecules, providing a high contrast between target and background tissues. NIR fluorophores have wider dynamic range and minimal background fluorescence as a result of reduced scattering compared with visible fluorescence detection. They also have high sensitivity, resulting from low background fluorescence, and high extinction coefficients, which provide high quantum yields. The NIR region is also compatible with solid-state optical components, such as diode lasers and silicon detectors. NIR fluorescence imaging is a noninvasive complement to radionuclide imaging in small animals or with probes in close proximity to the target in humans (4). Among the various optical imaging agents, only indocyanine green (
Sperm protein 17 (Sp17) is expressed exclusively in the testes, with its primary function of binding to the zona pellucida for fertilization (9). Expression of Sp17 in other normal tissues is limited. However, Sp17 was later found to be aberrantly expressed in various malignant tumors, such as myeloma, ovarian tumors, and esophageal squamous cell tumors (10, 11). Sp17 was expressed in 66% of endometrial tumors and 61% of cervical tumors (12). Li et al. (13) explored the overexpression of Sp17 in malignant tumors as a molecular biomarker for tumor imaging by coupling anti-human Sp17 monoclonal antibody with ICG derivative 02 (anti-Sp17-ICG-Der-02). ICG-Der-02 contains one carboxyl functional group for covalent conjugation to the amino group of biomolecules. ICG-Der-02 exhibits favorable water solubility and higher fluorescence quantum yield than ICG. Anti-Sp17-ICG-Der-02 was evaluated for in vivo NIR imaging of Sp17-positive human hepatocellular carcinoma cell line SMMC-7721 xenografts in mice.