A collection of fresh frozen hepatocellular carcinoma (HCC, n=38) samples was used in the
present study. HCC specimens were collected at the University of California, San Francisco (San Francisco, CA). Institutional
Review Board approval was obtained and informed consent was obtained from all subjects. See Gordan et al., manuscript
in preparation, for further details of this sample set. Immunofluorescence staining and imaging was performed at
the Gladstone Institutes Histology and Light Microscopy core using fluorescently-labeled primary antibodies (all from Cell
Signaling, Danvers, MA) to CTNNB1 (Mouse mAb L54E2, conjugated to Alexa Fluor® 555, 1:200, Cat#5612) YAP
(Rabbit mAb D8H1X, conjugated to Alexa Fluor® 488, 1:200, Cat#14729) and LKB1 (Rabbit mAb, 1:250,
Cat#13031, followed by goat anti-rabbit conjugated to Alexa Fluor® 633, 1:200). Staining and imaging for
DAPI/YAP/LKB1 was first performed on 38 samples, and 5 months later serial sections for 25 of these samples were
stained/imaged for DAPI/YAP/CTNNB1. Stitched images of entire sections were acquired at 20× (0.32μm per pixel)
on automated slide scanners (BZ-X700; Keyence, Osaka, Japan; Aperio VERSA Digital Pathology Scanner, Leica Biosystems, and
Axio Scan.Z1, Carl Zeiss Microscopy).
present study. HCC specimens were collected at the University of California, San Francisco (San Francisco, CA). Institutional
Review Board approval was obtained and informed consent was obtained from all subjects. See Gordan et al., manuscript
in preparation, for further details of this sample set. Immunofluorescence staining and imaging was performed at
the Gladstone Institutes Histology and Light Microscopy core using fluorescently-labeled primary antibodies (all from Cell
Signaling, Danvers, MA) to CTNNB1 (Mouse mAb L54E2, conjugated to Alexa Fluor® 555, 1:200, Cat#5612) YAP
(Rabbit mAb D8H1X, conjugated to Alexa Fluor® 488, 1:200, Cat#14729) and LKB1 (Rabbit mAb, 1:250,
Cat#13031, followed by goat anti-rabbit conjugated to Alexa Fluor® 633, 1:200). Staining and imaging for
DAPI/YAP/LKB1 was first performed on 38 samples, and 5 months later serial sections for 25 of these samples were
stained/imaged for DAPI/YAP/CTNNB1. Stitched images of entire sections were acquired at 20× (0.32μm per pixel)
on automated slide scanners (BZ-X700; Keyence, Osaka, Japan; Aperio VERSA Digital Pathology Scanner, Leica Biosystems, and
Axio Scan.Z1, Carl Zeiss Microscopy).