Figures 3C and 3D show examples of labelling 1 week and 2 Pevonedistat concentration weeks respectively; these both resemble the material at 1 hour survival. At survival times of 2 weeks or longer (Figure 3D), the fluorescent microspheres appeared somewhat larger than at shorter times, possibly indicating the microspheres were being sequestered together in phagosomes. Microspheres could be detected at survival times of 6 weeks, the longest time investigated in this study. Figure 3 Merged images of fluorescence photomicrographs from animals injected intravenously at P20 show Alexa 488

(green) labelled and large (0.2 μm) red fluorescent microsphere containing cells. A: 30 minutes following IV injection. B: 1 hr following injection. C: 1 week following injection. D: 2 weeks following injection. Calibration bar in ‘D’ = 50 μm for all images. Comparison of IP and IV injections One of the goals of this study was to determine the age at which Kupffer cells would show phagocytosis of fluorescent microspheres. Intravenous injections in younger mouse pups are challenging, so

the efficacy of intraperitonal (IP) injections this website was explored. Figure 4 compares microsphere labeling of liver cells from age matched animals, both injected with the larger 0.2 μm microspheres at P16. One received an intravenous (IV) tail vein OSI-906 supplier injection of fluorescent microspheres (Figure 4A,B,C) and the other (Figure 4D,E,F) RVX-208 receiving an IP injection. Both animals were euthanized 1 hour after the injection. The two injection procedures resulted in very similar distributions of labelling within the liver, with evidence of red fluorescent microspheres within green F4/80 immunoreactive

cells in both cases (Figure 4C,F). Although the distributions of the fluorescently labelled microspheres in the two experimental paradigms were virtually identical, the IV injections typically yielded more intense labelling (compare Figure 4A and 4D). Because the present study was not intended as a quantitative assessment of phagocytic uptake of markers but rather a study of cell types that accumulate the microspheres, these data were interpreted to indicate that an IP injection could be used with confidence when conducting experiments on the small early postnatal mice. Figure 4 Fluorescence images allow comparison of results of IV and IP injections. Fluorescence images under rhodamine optics show labelling of mouse liver 1 hr following intravenous (A) or intraperitoneal (D) injections of red labelled large (0.2 μm) microspheres. The same sections were photographed under fluorescein optics (B and E) to show F4/80 immunoreactivity. Merged images in C and F demonstrate co-localization of red microspheres and green immunoreactivity. Calibration bar in F = 50 μm for all images.