Biophot microscope

Immunohistochemical analysis was performed as previously described [41 (link),44 (link)]. Briefly, paraffin-embedded mouse liver tissues were sectioned, and the sections were placed on microscope slides. The deparaffinized sections were boiled in buffered sodium citrate solution (0.01 mol/L, pH 6.0) for 10 min and subjected to immunohistochemical staining with anti-Ki67 antibody (ab16667, 1:200; Abcam) and anti-GFP antibody (ab6673, 1:400; Abcam) followed by horseradish peroxidase-linked secondary antibody for 30 min. The tissues were stained with 3,3′-diaminobenzidine using the ImmPACT DAB (Vector Laboratories, Burlingame, CA, USA) or the ChemMate EnVision Kit (Agilent Technologies, Inc., Santa Clara, CA, USA). Slides were immersed in hematoxylin for counterstaining and then observed under a Nikon Biophot microscope (Japan) and photographed using a digital camera (Nikon Digital Sight DS-Ri1, Tokyo, Japan).

Immunohistochemistry was conducted as previously described [30 (link)]. Briefly, paraffin-embedded xenograft tumor tissues were sectioned and placed on glass microscope slides. After the sections were deparaffinized, they were boiled in buffered sodium citrate solution (0.01 mol/L, pH 6.0) for 10 min and subjected to immunohistochemical staining with anti-Ki-67 antibody (ab15580, 1:1000; Abcam, Cambridge, UK), followed by horseradish peroxidase-conjugated secondary antibody for rabbit IgG for 30 min. Then, the tissues were stained with 3,3′-diaminobenzidine using the ChemMate EnVision Kit (Agilent Technologies, Inc., Santa Clara, CA, USA). The slides were briefly immersed in hematoxylin for counterstaining and then observed under a Nikon Biophot microscope (Nikon, Tokyo, Japan) and photographed using a digital camera (Nikon Digital Sight DS-Ri1, Nikon). The photographs were taken under 400× magnification (Supplementary Figure S3A). Ki-67-positive cells were counted from five randomly selected fields (Supplementary Figure S3B).