Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: Immunohistochemical analysis of CD133 expression demonstrating the scoring intensity and expression pattern. Representative images using anti-CD133 MAb ab5558: ( A ) liver cholangiocarcinoma with multifocal, minimal to mild membranous and cytoplasmic staining, ( B ) pancreatic adenocarcinoma with mild to moderate membranous (apical) staining of luminal structures, ( C ) gastric adenocarcinoma with moderate to strong staining in two distinct cell populations: (1) luminal and apical and (2) cytoplasmic and membranous, ( D ) normal liver with minimal and nonspecific cytoplasmic staining of hepatocytes and apical staining of bile duct, ( E ) normal pancreas with weak to mild, apical membranous staining of acinar epithelium and ductal epithelium, ( F ) normal stomach with minimal to mild apical staining of glandular crypt epithelium. The scale bars represent 50 μ m.

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: Immunohistochemical staining, Expressing, Staining

Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: CD133 expression analysis by immunohistochemistry

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: Expressing, Immunohistochemistry

Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: Tumour and normal cell line expression of CD133 and sensitivity to anti-CD133 antibody-drug conjugate, AC133-vcMMAF

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: Expressing, Inhibition

Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: Activity of anti-CD133 ADC against cancer cell lines. ( A ) ADCs targeting CD133 have potent cytotoxic activity against antigen-positive hepatocellular and gastric carcinoma cell lines. Cytotoxicity was measured by resazurin dye conversion in Hep3B and KATO III cells grown in 96-well plates and exposed to anti-CD133 (AC133-vcMMAF) and control ADCs (IgG-vcMMAF and OKT9-vcMMAF) and crosslinked unconjugated anti-CD133 MAb (AC133) for 96 h. ( B ) Proliferation was measured using [ 3 H]-thymidine uptake in Hep3B and KATO III cells grown in 96-well plates and exposed to anti-CD133 and control ADCs for 96 h. ( C ) Induction of apoptosis in Hep3B cells treated with AC133-vcMMAF. Caspase 3/7 activation, a quantitative measurement of apoptotic cells, was monitored using the Caspase Glo assay at various time points (24–72 h) after addition of ADCs. Caspase 3/7 activation relative to untreated cells was detected by 48 h with optimal measurement after 72 h in Hep3B cells treated with increasing concentrations of AC133-vcMMAF and positive control OKT9-vcMMAF. ( D ) Inhibition of ADC cytotoxicity using internalisation inhibitor, ammonium chloride (NH 4 Cl) in Hep3B cells. Cells were incubated with increasing concentration of NH 4 Cl 30 min before treated with anti-CD133 (AC133-vcMMAF) or control ADCs. Cytotoxicity was measured after 72 h using the rezasurin dye conversion as in ( A ). The percentage inhibition of cytotoxicity relative to control untreated cells at an ADC concentration of 400 ng ml −1 is shown.

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: Activity Assay, Control, Activation Assay, Caspase-Glo Assay, Positive Control, Inhibition, Incubation, Concentration Assay

Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: Subcellular localisation of anti-CD133 ADC, AC133-vcMMAF, in sensitive and resistant cancer cell lines. ( A ) AC133-vcMMAF, partially colocalises (yellow) with the lysosomal marker, CD107a, in Hep3B and KATO III cells. Subcellular localisation of AC133-vcMMAF (red) and CD107a (green) in Hep3B and KATO III cells after 24 h incubation with the ADC. ( B ) Subcellular localisation of AC133-vcMMAF, lysosomal marker, CD107a, and caveolin-1 (Cav-1) in Su.86.86 after 24 h incubation with the ADC. AC133-vcMMAF colocalises with Cav-1 (yellow) and not with CD107a in this resistant cell line. Nuclei were stained blue with DAPI. Images were acquired using a × 63 oil immersion objective with Apotome for optical sectioning.

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: Marker, Incubation, Staining

Journal: British Journal of Cancer

Article Title: CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers

doi: 10.1038/sj.bjc.6604437

Figure Lengend Snippet: In vivo efficacy of an anti-CD133 ADC in Hep3B hepatocellular carcinoma model including IHC analysis of CD133 expression following ADC treatment. ( A ) In vivo efficacy of AC133-vcMMAF in Hep3B subcutaneous tumours. SCID mice ( n =7/group) with established (∼100 mm 3 ) Hep3B tumour xenografts were treated by intraperitoneal injection every 4 days for a total of four doses (red arrows) with the anti-CD133 antibody (AC133) or ADC (AC133-vcMMAF) or isotype control mouse IgG1-vcMMAF. An additional group of mice was left untreated as a control. Median tumour volume plots were continued for each group until one or more animals died or were euthanised (see Materials and methods). Tumours were collected when the tumour volume reached 1000 mm 3 . Highly concordant data were in an independent replicate of this experiment. ( B ) CD133 expression in Hep3B xenograft tumours after anti-CD133 drug conjugate treatment. IHC analysis using rabbit anti-CD133 MAb in ( a ) untreated Hep3B xenograft, ( b ) treatment with control IgG-vcMMAF (3.0 mg kg −1 ) and ( c ) treated with AC133-vcMMAF (3.0 mg kg −1 ). ( d ) same tumour as ( c ) stained with control rabbit IgG as primary antibody. Fast Red chromagen was used to detect CD133 expression.

Article Snippet: For mouse xenograft tumours, a rabbit anti-CD133 MAb (Cell Signaling Technology, Danvers, MA, USA) and a rabbit polyclonal anti-CD133 (ab19898, Abcam) were used separately as primary antibodies and the Bond Polymer Alkaline Phosphate (AP) Red Detection kit (Vision BioSystems, Australia) for detection.

Techniques: In Vivo, Expressing, Injection, Control, Staining

Journal: Analytical Cellular Pathology (Amsterdam)

Article Title: Effect of Shogaol on the Expression of Intestinal Stem Cell Markers in Experimentally Induced Colitis in BALB/c Mice

doi: 10.1155/2019/5134156

Figure Lengend Snippet: Cytoplasmic and membranous CD133 expression in epithelial cells of the colonic crypts. (a) Minimal, week expression (sum score 0) in mice of control negative group. (b) Focal, weak expression (sum score 1) in control positive group. (c) Focal, weak-moderate expression (sum score 4) deeply within the colonic crypt in group 3 (DSS exposure & 6-TG treatment). (d) Diffuse, weak expression (sum score 3) in group 4 (DSS exposure & 10 mg/kg b.w. Shogaol treatment). (e) and (f) Diffuse, moderate expression (sum score 6) extends into the transient amplifying and apical regions of the colonic crypts in group 5 (DSS exposure & 20 mg/kg b.w. Shogaol treatment) and group 6 (DSS exposure & 40 mg/kg b.w. Shogaol treatment), respectively.

Article Snippet: Following that, the slides were divided into 2 sets: the first set was incubated overnight at 4°C with rabbit anti-CD133 polyclonal Abs (1 : 100, San Francisco Biorbyt, USA), and the second set was incubated with rabbit anti-CD34 monoclonal Abs (1 : 100, Dako, Germany).

Techniques: Expressing, Control

Journal: Analytical Cellular Pathology (Amsterdam)

Article Title: Effect of Shogaol on the Expression of Intestinal Stem Cell Markers in Experimentally Induced Colitis in BALB/c Mice

doi: 10.1155/2019/5134156

Figure Lengend Snippet: Immunofluorescence expression of the CD133 protein in the cytoplasm of the colonic cryptal epithelium. (a) and (b) Weak (+) expression in mice of the control negative and control positive group. (c) and (d) Weak-moderate (+ to ++) expression in the in deep portions of the colonic crypts in mice of group 3 (6-TG treatment) and group 4 (10 mg/kg b.w. Shogaol treatment). (e) and (f) Moderate (++) expression in mice of group 5 (20 mg/kg b.w. Shogaol treatment) and mice of group 6 (40 mg/kg b.w. Shogaol treatment).

Article Snippet: Following that, the slides were divided into 2 sets: the first set was incubated overnight at 4°C with rabbit anti-CD133 polyclonal Abs (1 : 100, San Francisco Biorbyt, USA), and the second set was incubated with rabbit anti-CD34 monoclonal Abs (1 : 100, Dako, Germany).

Techniques: Immunofluorescence, Expressing, Control

Journal: PeerJ

Article Title: DNA mismatch repair and CD133-marked cancer stem cells in colorectal carcinoma

doi: 10.7717/peerj.5530

Figure Lengend Snippet: (A) The weak CD133 staining seen at the luminal border of some normal colonic crypts is considered equivocal and interpreted as negative (x200). In contrast, (B) is a case of colorectal carcinoma with strong CD133 staining of the glandular luminal surface (x200) and (C) a case of poorly differentiated colorectal carcinoma demonstrating CD133 staining as tiny cytoplasmic blobs which represent abortive glands (x200). (D) Colorectal carcinoma with deficient DNA mismatch repair (represented by loss of MLH1). Note the lack of immunoexpression of MLH1 in the colorectal carcinoma nuclei in the presence of immunopositivity in the internal positive controls (stromal cells, lymphocytes and nearby normal enterocytes) (x200)

Article Snippet: Immunohistochemical staining with a rabbit polyclonal antibody to CD133 (1:200; Abcam ab19898, Cambridge, United Kingdom) and detection by the ultraView universal DAB detection kit (Ventana Medical Systems Inc., Tucson, AZ, USA) was carried out on the Ventana Benchmark XT autostainer (Ventana Medical Systems Inc., Tucson, AZ, USA).

Techniques: Staining

Journal: PeerJ

Article Title: DNA mismatch repair and CD133-marked cancer stem cells in colorectal carcinoma

doi: 10.7717/peerj.5530

Figure Lengend Snippet: CD133 expression and DNA mismatch repair (MMR) status. The CD133 expression and MMR status as per size, TNM stage and differentiation of the colorectal carcinoma ( n = 80).

Article Snippet: Immunohistochemical staining with a rabbit polyclonal antibody to CD133 (1:200; Abcam ab19898, Cambridge, United Kingdom) and detection by the ultraView universal DAB detection kit (Ventana Medical Systems Inc., Tucson, AZ, USA) was carried out on the Ventana Benchmark XT autostainer (Ventana Medical Systems Inc., Tucson, AZ, USA).

Techniques: Expressing

Journal: PeerJ

Article Title: DNA mismatch repair and CD133-marked cancer stem cells in colorectal carcinoma

doi: 10.7717/peerj.5530

Figure Lengend Snippet: DNA mismatch repair protein (MMR) status and CD133 expression. CD133 immunoexpression and score being stratified by MMR status in right-sided and left-sided colorectal carcinomas.

Article Snippet: Immunohistochemical staining with a rabbit polyclonal antibody to CD133 (1:200; Abcam ab19898, Cambridge, United Kingdom) and detection by the ultraView universal DAB detection kit (Ventana Medical Systems Inc., Tucson, AZ, USA) was carried out on the Ventana Benchmark XT autostainer (Ventana Medical Systems Inc., Tucson, AZ, USA).

Techniques: Expressing