All animal experiments were conducted in accordance with University of Washington Institutional Animal Care and Use Committee (IACUC) approved protocols (IACUC 3441-05) as well as with federal guidelines. C57BL/6J WT mice (Jackson Laboratories) were systemically administrated via tail vein injection with Dy677 labeled NP-siScrambled-CTX (NP-siScr-Dy677-CTX as a siRNA control NP). The non-injected animal was included in the study as control. Two and or forty-eight hours after injection, the mice were euthanized and tissues were dissected from brain, liver, kidneys and spleen. Each tissue was imaged using a Xenogen IVIS Lumina II system (Perkin Elmer) to examine NP distribution.
Blood was collected by retro-orbital eye bleed or terminal heart puncture for blood half-life evaluation at 0.5, 1, 2, 4, 24, and 48 hr after NP injection. Due to the limited amount of blood from each animal, none were used for more than two time points. Blood samples were drawn from four independent mice for each time point. Blood (100 μl) was added to a 96 well clear bottom plate and scanned on the Odyssey NIR fluorescence imaging instrument (LI-COR) using the 700 nm-channel (λexc = 685 nm with λem = 705 nm). The concentration of NP was interpolated from the NP-siScr-Dy677-CTX fluorescence standard curve.
Blood was collected by retro-orbital eye bleed or terminal heart puncture for blood half-life evaluation at 0.5, 1, 2, 4, 24, and 48 hr after NP injection. Due to the limited amount of blood from each animal, none were used for more than two time points. Blood samples were drawn from four independent mice for each time point. Blood (100 μl) was added to a 96 well clear bottom plate and scanned on the Odyssey NIR fluorescence imaging instrument (LI-COR) using the 700 nm-channel (λexc = 685 nm with λem = 705 nm). The concentration of NP was interpolated from the NP-siScr-Dy677-CTX fluorescence standard curve.