IHC and FISH analyses of HAT mice. One day after normal saline (control) and acetaminophen administration, each mouse received a tail vein injection of donor cells. Two months after transplantation, mice were killed and liver cryosections were analyzed by IHC; 5-μm serial sections were stained with anti-GFP, antialbumin, anti-CK18, and anti-vWF antibodies. (A) Normal saline-injected mice. The merged picture shows that eGFP-expressing cells did not engraft in any of the sections examined. The sections were stained positive for albumin, CK-18, and vWF expressed by the host tissue (n = 4). (B) Acetaminophen-injected mice. The merged picture shows that donor-derived (eGFP⁺) cells were present in all the sections examined and expressed albumin (top panel), CK-18 (middle panel), and vWF (bottom panel; n = 4). (Insets) Magnified region of the tissue sections. As secondary antibody controls, the sections were also stained with respective conjugates (data not shown). (C) Sex chromosome FISH and immunocytochemistry. Liver cells of HAT mice (10 months of transplantation) were labeled with X (fluorescein isothiocyanate) and Y (Cy3) chromosome-specific probes. The fusions between donor (XX) and recipient (XY) cells were microscopically examined. White arrows represent donor-derived unfused cells (left). The same liver cells were immunostained for albumin and CK-18. The merge pictures show that donor-derived (eGFP⁺) cells express both albumin (middle) and CK-18 (right), marked by white arrows. Original magnifications: albumin and CK-18, ×20; vWF, ×40; and FISH, ×100.