Fig. 6.

In vivo pharmacokinetics and ex vivo high resolution NIRF imaging (tumor sections) of the 5D3 IgG and Fab fragment. Panels A–F: A mouse was co-injected with the IgG-IRDye800CW (A) and the Fab-IRDye680RD (B) with overlay shown in (C), where IgG is displayed in green and the Fab is displayed in red. In (A), high tumor contrast is achieved 12 hr post-injection and by 24 hr the whole-mouse background uptake is also low. In (B), high tumor contrast is achieved already 2 hr post-injection [gastrointestinal (GI) signal is chlorophyll] and continues till 48 hr post-injection. The overlay in row (C) shows a high degree of co-localization from 12 hr onwards. Panels (D, E) show the 72 hr uptake without skin of the IgG and Fab, respectively. Both are selective for the PSMA-positive tumor at 72 hr. Panel (F) reveals largely yellow co-localization of both antibody formats within the PSMA-positive tumor. Panels G, H: PSMA-positive PC-3 PIP and PSMA-negative PC-3 flu tumors were harvested following imaging, sectioned and scanned to detect the high-resolution distribution of IgG-IRDye800CW and Fab-IRDye680RD within the tumors. Row (G) shows a section of the PSMA-positive PC-3 PIP tumor where green depicts IgG, red depicts Fab and yellow shows co-localized IgG and Fab in the leftmost section. Row (H) shows the same investigation for sections of PSMA-negative PC-3 flu tumor (dotted lines). In the PSMA-positive section, IgG uptake (green) appears more confined to the tumor rim and small focal regions near the rim while the Fab (red) appears to display a wider uptake pattern both away from the rim and within tumor.