Figure 4c:

(a) Mechanism of action for dual-modality, activatable, magneto-optical molecular imaging MR contrast agent, which consists of a core of a dextran-coated iron oxide nanoparticle, onto which fluorochrome A (small purple spheres) is conjugated. Also attached to its surface are peptides carrying a second fluorochrome B (blue spheres) that can be cleaved by a specific enzyme. When the contrast agent enters an environment where the specific enzyme is present (eg, cathepsin B, which is overexpressed in breast cancer), the fluorochrome-carrying peptides are cleaved and thus fluorochrome B detaches. The increased distance of the fluorochromes causes dequenching, resulting in increased fluorescence signal. The intensity of fluorochrome A remains the same. (b–d) Illustration of fluorescence ratio method. Fluorescence is normally depth dependent and difficult to quantify. However, if two fluorochromes are used, where one is nonactivatable and serves as an internal standard, correction for depth is possible. While the signal from both fluorochromes used here (Cy5.5 and Cy7) decreases with increasing depth, the ratio of the two signal intensities remains the same (c, d). (Images b–d reprinted, with permission, from reference 33.)