Rabbit anti ki67

Immunostaining was performed in paraffin sections according to the standard protocols. In brief, samples were deparaffinized and rehydrated, followed by antigen retrieval in 10 mm sodium citrate (Beyotime, P0083). Blocking and staining were performed in antibody diluent with 10% Goat serum. Sections were incubated with mouse anti‐Pdgfrα (Santa Cruz, sc‐398206) and rabbit anti‐Ki67 (Bethyl Laboratories, IHC‐00375) antibodies overnight at 4 °C, followed by incubation with the corresponding secondary antibodies goat anti‐mouse Alexa Fluor 594 (Thermo Fisher, A11032) and anti‐rabbit Alexa Fluor 488 (Thermo Fisher, A11034), respectively. Slides were mounted with DAPI (4′,6‐diamidino‐2‐phenylindole) and imaged by fluorescence microscopy. Cell counting was achieved using the ImageJ software.

Tissue samples were collected from WT and L22P mice, washed in Phosphate-buffered saline (PBS), and fixed in 10% neutral-buffered formalin solution (pH 7.4) overnight. Tissues were embedded in paraffin and serially sectioned by the MD Anderson Research Histology Core Facility. Immunostaining for Ki67 and γH2AX was performed using rabbit anti-Ki67 (Bethyl Laboratories, Montgomery, TX, USA IHC-00375-1) and rabbit anti-γH2AX (Cell Signaling, Danvers, MA, USA 2577), respectively. Samples were imaged at 40× magnification with bright field using Aperio Scanscope (Nikon Instruments Inc., Melville, NY, USA), Leica Biosystems (Buffalo Grove, IL, USA). Data for Ki67 and γH2AX were scored blindly using two independent lab individuals. To score Ki67 and γH2AX-positive cells counted using each tissue section, at least 10 fields were used, and the average percentage of Ki67-and γH2AX-positive cells in three mice for every condition was plotted. Immunohistochemical analysis was performed as described previously [29 (link)]. Histopathological scoring of gastric mucosa was performed as described previously [30 (link),31 (link)].