codex staining  (Akoya Biosciences)

Journal: Cancer Discovery

Article Title: Stromal KITL/SCF Maintains Pancreas Tissue Homeostasis and Restrains Tumor Progression

doi: 10.1158/2159-8290.CD-24-1079

Figure Lengend Snippet: KITL regulates PSC state and pancreas tissue homeostasis. A, Volcano plot of all upregulated, nonsignificant, and downregulated differentially expressed genes (DEG) as defined by the Wald test ( P .adj < 0.05 and log 2 FC > 1) from Kitl overexpression ( Kitl OE) ImPSC-1 bulk RNA-seq dataset with representative gene labels included. Data represent three biological repeats. B, Gene ontology (GO) analysis of upregulated and downregulated genes in immortalized PSCs (ImPSC-1) overexpressing Kitl . Top 10 enrichment categories ranked by adjusted P values plotted in each direction. C, qRT-PCR analysis of indicated DEGs in ImPSC-1 expressing stable Kitl knockdown (sh Kitl ) or overexpression ( Kitl OE). PLKO vector (shCtrl) and fluorescence overexpression ( Egfp OE) serves as the respective control. D, Representative images of CODEX staining (left) and quantification (right) for GFP (green) and CD31 (red) in normal pancreas from Fabp4-Cre or Kitl fl/fl ;Fabp4-Cre mouse model ( n = 3–4 mice per arm). Scale bar, 100 μm. E, Representative images of CODEX composite staining (left) and quantification (right) for CD45 (white) in normal pancreas from Fabp4-Cre or Kitl fl/fl ;Fabp4-Cre mouse model ( n = 4 mice per arm). Scale bar, 50 μm. F, Representative images of IHC staining for GFP (green), c-KIT receptor (KIT, red), and VIM (magenta) in healthy murine pancreas from Rosa26 mTmG/+ ;Fabp4-Cre mice. Scale bar, 10 μm. G, Representative images of IHC staining for CD31 (green) and c-KIT receptor (KIT, red) in healthy murine pancreas. Scale bar, 10 μm. H, Representative images of IHC staining for panCK (white) and c-KIT receptor (KIT, red) in healthy murine pancreas. Scale bar, 10 μm. I, Representative hematoxylin and eosin images between caerulein-treated Fabp4-Cre and Kitl fl/fl ;Fabp4-Cre mice. Scale bar, 100 μm. J, Representative images of IHC staining (left) for panCK (green) and amylase (red) between caerulein-treated Fabp4-Cre and Kitl fl/fl ;Fabp4-Cre mice. Scale bar, 10 μm. PanCK quantification on right ( n = 3 mice per arm). K, Representative IHC images (left) and quantification (right) for CD45 staining between caerulein-treated Fabp4-Cre and Kitl fl/fl ;Fabp4-Cre mice. Scale bar, 100 μm. L, Representative images (left) and quantification (right) for CD68 staining in caerulein-treated Fabp4-Cre and Kitl fl/fl ;Fabp4-Cre mouse pancreas. The same co-stained tissue section is shown in K and L . Scale bar, 100 μm. For comparisons of two groups, an unpaired t test was used. For comparisons of more than two groups with one independent variable, the one-way ANOVA test was used. Data are represented as mean ± SEM; *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001; FC, fold changes; ns, not significant; panc area, pancreas.

Article Snippet: In brief, 5-μm FFPE pancreas sections were mounted onto 22 × 22 mm glass coverslips (Electron Microscopy Sciences) coated in 0.1% poly-L-lysine (Sigma-Aldrich) and stained with using CODEX Staining Kit (Akoya Biosciences).

Techniques: Over Expression, RNA Sequencing, Quantitative RT-PCR, Expressing, Knockdown, Plasmid Preparation, Fluorescence, Control, Staining, Immunohistochemistry