Apc anti mouse cd326 epcam

An adult Rosa^YFP/YFP mouse received retro-orbital injection of AAV-TBG-cre at the dose of 2.5 × 10¹¹ viral particles, and hepatocytes were harvested 3 weeks later using a standard two-step collagenase perfusion method. Isolated Rosa^YFP/YFP hepatocytes were stained with APC anti-mouse CD326/EPCAM) (1:100, G8.8, Biolegend) or APC anti-human/mouse CD49f/Itga6 antibody (1/100, GoH3, Biolegend). For staining with Prom1/Cd133 and Cd24, cells were incubated with purified rat anti-mouse CD133/Prom1 antibody (1:100, 315–2 C11, Biolgend) and rat anti-mouse CD24 antibody (1:100, Biolegend, M1/69) followed by staining with Alexa647-conjugated donkey anti-rat antibody (1:300, Jackson ImmunoResearch). APC-conjugated rat IgG2a or IgG2b, κ isotype control antibody was used as control. DAPI was added to stain dead cells. Attune NxT Flow Cytometer (Lifetechnologies) was used for data collection. B6 wild-type adult hepatocytes, which were stained with only DAPI, was used for making the threshold of YFP signal.

For flow cytometry, PE–Cy5 anti-NK1.1 (5 μl per 100 μl of sample, clone: PK136), FITC anti-CD3 (5 μl per 100 μl of sample, clone: 17A2), APC anti-CD3 (5 μl per 100 μl of sample, clone: 17A2), FITC anti-DX5 (5 μl per 100 μl of sample, clone: DX5), FITC anti-CD4 (5 μl per 100 μl of sample, clone: RM4-5), APC anti-CD8 (5 μl per 100 μl of sample, clone: SK1), FITC anti-γδ TCR (5 μl per 100 μl of sample, clone: GL3), APC anti-Ly49A (5 μl per 100 μl of sample, clone: A1/Ly49A), PE anti-NKG2A (5 μl per 100 μl of sample, clone: 16A11), PE anti-NKG2D (5 μl per 100 μl of sample, clone: 1D11), APC anti-KLRG1 (5 μl per 100 μl of sample, clone: 2F1/KLRG1), PE anti-IL-22 (5 μl per 100 μl of sample, clone: Poly5164), APC anti-IL-22 (5 μl per 100 μl of sample, clone: Poly5164), PE anti-IFN-γ (5 μl per 100 μl of sample, clone: XMG1.2), APC anti-CD49a (5 μl per 100 μl of sample, clone: HMα1), APC anti-mouse CD326 (Ep-CAM) (5 μl per 100 μl of sample, clone: G8.8) and APC anti-CXCR6 (5 μl per 100 μl of sample, clone: K041E5) (all from BioLegend), Anti- mouse IL-22R alpha 1 (2.5 μg per 10⁶ cells, clone: 496514), Rat IgG2A Isotype Control (2.5 μg 10⁻⁶ cells, clone: 54447) and Anti mouse Proinsulin (2.5 μg per 10⁶ cells, clone: 253627) (from R&D systems) were used.