Journal: Advanced Science

Article Title: Colorectal Cancer Stem Cell States Uncovered by Simultaneous Single‐Cell Analysis of Transcriptome and Telomeres

doi: 10.1002/advs.202004320

Figure Lengend Snippet: Validation of enriched genes shared by CSCs and EPCs or specific for CSCs by immunofluorescence and telomere FISH of CRCs. a) Proliferation markers PCNA or MKI67 and shared markers KRT18, SLC12A2, CXADR, and CLDN4. b) Proliferation marker MKI67 costained with CSC enriched genes, TNFRSF19 and BAMBI. White arrows indicate marker positive but MKI67 negative cells. c) CSC specific markers TNFRSF19 or BAMBI (green) and telomere FISH (red). Areas within dashed lines in orange indicate marker positive cells; areas within dashed lines in white indicate marker negative cells. d) CSC and EPC shared markers SLC12A2 or KRT18 and telomere FISH (red). Areas within dashed lines in orange indicate marker positive cells; areas within dashed lines in white indicate marker negative cells. Scale bar = 20 µm.

Article Snippet: After being deparaffinized, rehydrated and washed in PBS, paraffin sections were incubated with 3% H 2 O 2 for 10 min at room temperature, subjected to high pressure antigen recovery sequentially in 0.01 m citrate buffer for 3 min, permeabilized in 0.1% Triton X‐100 for 30 min, blocked with 5% goat serum with 0.1% BSA in PBS for 2 h at room temperature, and then incubated with the primary antibodies against MKI67 (AB9260, Millipore), PCNA (sc‐25280, Santa Cruz), E‐CADHERIN (20874‐1‐AP, Proteintech), TNFRSF19 (sc‐398526, Santa Cruz), BAMBI (sc‐100681, Santa Cruz) KRT18 (A1022, ABclonal), CLDN4 (sc‐376643, Santa Cruz), CXADR (sc‐373791, Santa Cruz), or SLC12A2 (sc‐514858, Santa Cruz) overnight at 4 °C.

Techniques: Biomarker Discovery, Immunofluorescence, Marker