Ethical approval was obtained from the Institutional Animal Care and Use Committee of Hoshi University (approval no. P22-016). A total of six female BALB/c mice (weight, 18–20 g; age, 8 weeks; Sankyo Labo Service Corporation, Inc.) were housed at 24°C and 55% humidity under a 12/12-h light/dark cycle (lights on at 8:00 a.m.) with ad libitum access to food and water. siRNA lipoplexes with 20 µg of Cy5-siRNA were systemically administered to mice via the lateral tail vein (n=1/siRNA lipoplex). At 1 h after injecting the siRNA lipoplexes, the mice were euthanized by cervical dislocation, and death was confirmed by cessation of the heartbeat. Tissues (lungs, heart, liver, spleen and kidneys) were analyzed through Cy5 fluorescence imaging using the NightOWL LB981 NC100 system (Berthold Technologies GmbH & Co. KG) as previously described (9 (link),13 (link)). Thereafter, the tissue samples were frozen on dry ice and sliced into 16-µm sections. The localization of the Cy5-siRNA was examined using a fluorescence microscope (Eclipse TS100-F; Nikon Corporation).