# Targeted Molecular Imaging of Pancreatic Cancer with a Miniature Endoscope
## 3 Results and Discussion  
Our previous study [33] showed that the light absorbance of IONPs increases with decreased wavelength; however, considering the fact that tissue penetration depth is greater in the NIR region, light with a wavelength of 730 nm was chosen as the wavelength of the light source for inducing photoacoustic signals in our experiments. Figure 4A-C, respectively, show photoacoustic MAP images from the mice of Group 1 administered with non-targeted nanoparticles NIR830-IONP, targeted NIR830-ATF-IONP, and PEG-coated targeted NIR830-ATF-PEG-IONP. To closely investigate these images, in Figure 4A, we can see that the contrast of tumor to normal tissue is too low to identify the tumor with non-targeted NIR830-IONP. However, for the tumor with targeted NIR830-ATF-IONP or PEG-coated targeted NIR830-ATF-PEG-IONP, greatly enhanced contrast of tumor to normal tissue can be seen in Figure 4B,C, where the tumor area with a clear tumor boundary is pinpointed. In comparing Figure 4B,C, it can be seen that the PEG coating deteriorates the contrast of the tumor to the background, but increases the specificity of tumor detection with a more clearly defined tumor boundary, since PEG was used to stabilize the nanoparticles and to modify surface properties to reduce the non-specific uptake of nanoparticles by macrophages in the reticuloendothelial system. We then selected a region of interest (ROI) within the tumor area, and computed the signal to background ratio (SBR). We plotted the SBR for the mice that had received the injection with three different nanoparticles, as shown in Figure 4D, for quantitative analysis. From these plots, we can see that the mouse with targeted NIR830-ATF-IONP has the highest SBR (\(\sim\)145, 43 dB), while the value for the mouse with non-targeted NIR830-IONP is the lowest (\(\sim\)24, 28 dB). PEG coating reduces the uptake of nanoparticles by cells like macrophages within the tumor, which is indicated by the contrast of the mouse with targeted NIR830-ATF-PEG-IONP (\(\sim\)102, 40 dB). After the endoscopic photoacoustic imaging, we then conducted experiments using the NIR planar fluorescence imaging system. Figure 5A-C show the fluorescence images from the mice of Group 1 injected with NIR830-IONP, NIR830-ATF-IONP, and NIR830-ATF-PEG-IONP, respectively. The highest contrast in the image for the mouse with non-targeted NIR830-IONP (Figure 5A) comes from both the tumor (indicated by red arrow) and normal tissue (indicated by blue arrow). Thus, the specificity is too low to identify the tumor. On the contrary, for the mouse with the targeted agent of NIR830-ATF-IONP (Figure 5B) or NIR830-ATF-PEG-IONP (Figure 5C), greatly enhanced image contrast was observed, allowing the tumor and the tumor boundary to be identified clearly. Taking a close look at the difference between the images for NIR830-ATF-IONP and NIR830-ATF-PEG-IONP, we note that the PEG coating reduces tissue uptake of nanoparticles, especially in organs without tumors, as indicated by the white arrows in Figure 5B.C. We also plotted the SBR of the ROI from these three mice, as shown in Figure 5D, where we see SBR values similar to those of the photoacoustic images (Figure 4D): The mouse with NIR830-IONP has a SBR of \(\sim\)35 (31 dB), while the mice with NIR830-ATF-IONP or NIR830-ATF-PEG-IONP, respectively, have an SBR of \(\sim\)144 (43 dB) and \(\sim\)101 (40 dB). Figure 4: Endoscopic photoacoustic images of pancreatic tumor from the mice of Group 1. Maximum amplitude projection (MAP) images of (**A**) mouse injected with NIR830-IONP; (**B**) mouse injected with NIR830-ATF-IONP; and (**C**) mouse injected with NIR830-ATF-PEG-IONP; (**D**) quantitative plot and comparison of average contrast in region of interest (ROI) (pancreatic tumor). The red scale bars indicate 3 mm in length. NIR, near-infrared; IONP, iron oxide nanoparticle; ATF, amino-terminal fragment; PEG, polyethylene glycol. Figure 5: Fluorescence images of pancreatic tumor from the mice of Group 1.