Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 1. The expression of CD133 in lung cancer and normal lung tissues. Original values are presented as log2 ratios. Spots showing the level of CD133 mRNA in NSCLC and normal tissues. (A) GSE10072. (B) GSE40275. (C) GSE63459; *p<0.05, **p<0.01 and ***p<0.001. (D) Correlation between CD133 and CXCR4 in GSE30219.

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: Expressing

Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 2. CD133 and CXCR4 are highly expressed in NSCLC patients with metastasis. (A) Representative images of immunohistochemistry staining (mag- nification, x400) of NSCLC patients. (a) CD133 low expression; (b) CD133 high expression; (c) CXCR4 low expression; (d) CXCR4 high expression. Positive staining ratio of CD133 (B)/CXCR4 (C) in metastatic and non-metastatic NSCLC patients. Positive staining score of CD133 (D)/CXCR4 (E) in metastatic and non-metastatic NSCLC patients. (F) Correlation results of CD133 and CXCR4. (G) Correlation analysis between CD133/CXCR4 co-expression and disease- free survival (red and black curves indicate high and low CD133/CXCR4 co-expression groups of patient death, respectively; *p<0.05).

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: Immunohistochemistry, Staining, Expressing

Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 3. CD133 enhances A549 cells proliferation. (A) Effectiveness of magnetic cell sorting was verified by immunofluorescence assay (magnification, x400). Red and blue indicate CD133 expression and cell nucleus, respectively. (B) Formation of colonies by A549 cell lines after 2-week incubation. (C and D) Cell proliferation capacity was determined in A549 cells, CD133+ A549 cells and CD133-A549 cells by CCK8 assays.

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: FACS, Immunofluorescence, Expressing, Incubation

Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 4. CXCR4 is upregulated by CD133. Silencing effectiveness of CD133 siRNA was verified by qPCR (A) and western blot analysis (B). (C) Expression of CXCR4 after treatment with CD133 siRNA (50 nM, 48 h) by qPCR analysis. (D) Western blot analysis for the expression of CXCR4 after treatment with CD133 siRNA (50 nM, 72 h). β-actin as a loading control. (E) The statistics of (D). All experiments were performed in triplicate. ***p<0.001.

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: Western Blot, Expressing, Control

Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 5. CD133+CXCR4+ promotes EMT process in NSCLC cells. (A) Transwell assay for the invasion of CD133-CXCR4+, CD133+CXCR4- and CD133+CXCR4+ cells. (B) Western blot assay for the expression of E-cadherin and Vimentin in CD133-CXCR4+, CD133+CXCR4- and CD133+CXCR4+ cells. (C) qPCR for the expression of E-cadherin, Vimentin, Snail, Slug and Twist in A549/CD133+, A549/CD133+ shRNA-NC, A549/CD133+ shRNA-CXCR4 and A549/CD133+ with amd3100. (D and E) RT-PCR for the expression of E-cadherin, Vimentin, Snail, Slug and Twist in A549/CD133+, A549/CD133+ shRNA-NC and A549/CD133+

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: Transwell Assay, Western Blot, Expressing, shRNA, Reverse Transcription Polymerase Chain Reaction

Journal: International journal of oncology

Article Title: CXCR4 is involved in CD133-induced EMT in non-small cell lung cancer.

doi: 10.3892/ijo.2016.3812

Figure Lengend Snippet: Figure 6. Vimentin is positively associated with CD133/CXCR4 co-expression. (A) Representative images of immunohistochemistry staining (magnification, x400) of NSCLC patients. (a) E-cadherin low expression; (b) E-cadherin high expression; (c) Vimentin low expression; (d) Vimentin high expression. Positive staining ratio of E-cadherin (B)/Vimentin (C) in metastatic and non-metastatic NSCLC patients. Correlation analysis for E-cadherin (D) or Vimentin (E) with CD133/CXCR4 co-expression in NSCLC patients.

Article Snippet: Cells were incubated at 4 ̊C with 100 μl FcR blocking reagent and 100 μl CD133 Micro Beads (130-097-049, Miltenyi Biotec, Germany) for 30 min.

Techniques: Expressing, Immunohistochemistry, Staining

Journal: Genes to Cells

Article Title: Functional Role of COP1 Gene in Hepatocellular Carcinoma Lipid Metabolism and Stemness

doi: 10.1111/gtc.70108

Figure Lengend Snippet: COP1 knockdown reduces motility, and stemness in HCC cells. (a‐b) Migration (a) or invasion (b) assay showing reduced motility in Huh7 and HepG2 cells treated with COP1‐siRNA, as quantified by the relative migration or invaded area. Statistical significance: *, p < 0.05; ** p < 0.01; *** p < 0.001 vs. control. (c) Sphere formation assay images and quantitative data from Huh7 and HepG2 cells, demonstrating a decrease in both the number and size of spheres in COP1‐siRNA‐treated cells. (d) Sphere formation assay images and quantitative data from PLC/PRF/5 CD133+ cells. Compared to the control grpup, both the sphere number and size decreased upon COP1‐siRNA treatment. Scale bar, 100 μm Statistical significance: *, p < 0.05; *** p < 0.001 vs. control.

Article Snippet: The cells were treated with FcR Blocking Reagent and CD133 MicroBeads (130‐100‐857, Miltenyi Biotec, Bergisch Gladbach, Germany) according to the manufacturer's instructions and incubated for 15 min at 4°C in the dark.

Techniques: Knockdown, Migration, Control, Tube Formation Assay

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: CD133 has upregulated expression in colorectal cancer tissues. (A) The HColA-180Su11 microarray (Outdo Biotech, Shanghai, China) containing 76 primary colon cancer samples was stained with an anti-CD133 antibody to determine the expression of CD133. Left: paracarcinoma tissue with negative CD133 expression; middle: primary colon cancer samples with lower CD133 expression; right: primary colon cancer samples with higher CD133 expression. Representative images (magnification ×4) are shown for CD133 immunostaining. The scale bar represents 500 μm. (B) The percentage of different levels of CD133 expression is indicated. (C) CD133 expression on colon cancer cell lines was analyzed by flow cytometry. (D) The HOrgN090PT02 microarray (Outdo Biotech) containing 25 human normal tissue samples was immunostained with an anti-CD133 antibody to determine the expression of CD133; human normal tissues are negative for CD133. Representative staining image fields (magnification ×20) are shown. Scale bars represent 100 μm.

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Expressing, Microarray, Staining, Immunostaining, Flow Cytometry

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: CAR133-NK92 cells with inducible secretion of CBLB502 enable superior proliferation, activation, and antitumor activity. (A) The absolute number of CAR133-NK92 and CAR133-i502-NK92 cells was analyzed when the cells were stimulated with CD133+ and CD133− antigens for different lengths of time (days). Data represent the mean ± SD of quadruplicate wells, * P < 0.05; NS, not significant. (B) The activation marker, CD69, was quantified by flow cytometry when CAR133-NK92 or CAR133-i502-NK92 cells were cultured with SW620 and SW480 cells for 24 h. The mean values from triplicate cultures are shown. *** P < 0.001. (C) Effector cells (NK92, CAR133-NK92, and CAR133-i502-NK92 cells) were co-cultured with colorectal cancer cells at an E:T ratio of 10:1 for 24 h. Effector cells were then harvested and CD107a expression was analyzed by flow cytometry. The mean values from triplicate cultures are shown. * P < 0.05; ** P < 0.01; NS, not significant. (D) Cell killing by CAR133-NK92 and CAR133-i502-NK92 cells was investigated by the LDH cytotoxicity assay after co-culture with target cells at the indicated E:T ratios. All data are expressed as the means ± SEMs of triplicate samples. * P < 0.05; NS, not significant. (E) The cytotoxicity of CAR133-NK92 and CAR133-i502-NK92 cells against colon cancer cells was calculated by RTCA. (F) CAR133-NK92 and CAR133-i502-NK92 cells were co-cultured with SW620 cells for 5 days, and the cytokines and chemokines in the culture supernatant were determined by the human cytokine array C5 (Ray Biotech, Atlanta, GA, USA). * P < 0.05. (G) The concentrations of GM-CSF and IL-7 in the supernatant were further determined by specific ELISA kits. Data represent the mean+SEM. * P < 0.05; ** P < 0.01.

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Activation Assay, Activity Assay, Marker, Flow Cytometry, Cell Culture, Expressing, LDH Cytotoxicity Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: CBLB502 promotes the antitumor activity of CAR133-NK92 cells in mice with subcutaneous CD133-expressing colon tumors. (A) Schedule of the challenge protocol. Mice were inoculated with CD133+ SW620 cells (1 × 10 6 cells/mouse) followed 14 days later by CAR133-i502-NK92, CAR133 NK92, or NK92 cells (5 × 10 6 cells/mouse, n = 5/group). (B) Colon cancer growth was monitored by in vivo bioluminescence imaging. (C) Kaplan–Meier survival curves showing the tumor-free survival of tumor-bearing mice treated with CAR133-i502-NK92, CAR133-NK92, or NK92 cells. Significance was determined by the log-rank test. *** P < 0.001. (D) Blood collected on day 15 was analyzed by flow cytometry for the presence of effector cells, CAR133-i502-NK92 cells showed enhanced proliferation in vivo . *** P < 0.001. (E) The number of CD56+ cells, representing NK92 cells, was increased in tumor samples of mice treated with CAR133-i502-NK92 cells compared to CAR133-NK92 and NK92 cells on day 15 after effector cell treatment. Each scale bar represents 50 μm. (F) Absolute number of CD56+ cells in . Data represent the mean ± SD of quadruplicate wells. *** P < 0.001.

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Activity Assay, Expressing, In Vivo, Imaging, Flow Cytometry

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: On-target off-tumor toxicities in the mouse model treated with CAR133-i502-NK92 cells. (A) The expression of mouse CD133 protein was tested in mouse stomach tissue using IHC assays. Representative staining image fields (magnification ×200) are shown. Scale bars represent 100 μm. (B) Histopathologic analysis of mouse organ tissues by H&E staining. Magnification ×20. Each scale bar represents 500 μm. (C) Detection of CAR-NK92 cells infiltrated into stomach tissues on day 15 after effector cell treatment by IHC staining of CD56 showed that there was minimal immune cell infiltration into the stomach (magnification ×100), The scale bar represents 200 μm. (D) Body weight of the individual mice from each treatment group. Data are presented as the mean ± SD of three independent experiments. Error bars represent the standard deviation.

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Expressing, Staining, Immunohistochemistry, Standard Deviation

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: Adoptively transferred CAR133-i502-NK92 cells successfully control tumor development in mice with CD133+ and CD133− mixed colon tumor models. (A) Schedule of the in vivo experimental design using CD133+ and CD133- mixed colon tumor model. (B) BLI illustrating colon cancer growth. (C) Kaplan–Meier survival curves generated from the survival of mice treated with CAR133-i502-NK92 and CAR133-NK92 cells. *** P < 0.001. (D) Expression of CD31 protein was tested in the mouse tumor tissue using IHC assays. Representative photomicrographs are shown (magnification ×400). The scale bar is 50 μm. *** P < 0.001. (E) Representative immunostaining images of CD3+ T-cell infiltration into cancer tissues. Representative staining of tumor sections in each experimental group is shown (magnification ×400). The scale bar represents 50 μm. *** P < 0.001.

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Control, In Vivo, Generated, Expressing, Immunostaining, Staining

Journal: Cancer Biology & Medicine

Article Title: The inducible secreting TLR5 agonist, CBLB502, enhances the anti-tumor activity of CAR133-NK92 cells in colorectal cancer

doi: 10.20892/j.issn.2095-3941.2023.0033

Figure Lengend Snippet: The schematic diagram of the anti-tumor activity of CAR133-NK92 cells in colorectal cancer. We engineered CAR133-NK92 cells with inducible secretion of CBLB502 and demonstrated that CAR133-i502-NK92 cells not only specifically eliminate CD133-positive colon cancer cells in a CAR133-dependent manner, but also indirectly eradicate CD133-negative colon cancer cells in a CBLB502-specific endogenous immune response manner. The figure was created with Biorender.com .

Article Snippet: To evaluate the surface expression of CD133 in colon cancer cell lines (SW620, SW480, and HCT116), the cells were washed with 1X PBS containing 4% BSA and incubated with PE-conjugated anti-CD133 antibody (ab253271; Abcam); IgG isotype controls were included.

Techniques: Activity Assay

Journal: PLoS ONE

Article Title: Reduced variability of neural progenitor cells and improved purity of neuronal cultures using magnetic activated cell sorting

doi: 10.1371/journal.pone.0213374

Figure Lengend Snippet: (A) Representative immunofluorescence images from one NPC line demonstrating a reduction in CD271+ cells and an enrichment of CD133+ positive cells following both FACS and MACS compared to unsorted cells. Cell nuclei are labelled with DAPI. Scale bar = 100μm, N = 6. (B) Combined flow cytometry analysis of percent CD271+, CD271- and CD271-/CD133+ cells from 100% live cells (DAPI negative population) comparing unsorted, FACS and MACS conditions. (C) Flow cytometry analysis in B presented as individual cell lines. (D) Reduction in CD271+ cell variability following MACS compared to unsorted cell s . **p<0.01 Variance F-test. (E) Fold change expression of stress-associated genes in cells following FACS compared to MACS. Dotted line denotes equal expression at fold change = 1. (F) Quantification of the percentage surface area covered by live cells imaged in D . (G) Absorbance read at 450nm of LDH assay carried out on media supernatant collected from cells 24 hours following either FACS or MACS sorting. (H) Cell viability assessed by Calcein Violet staining 24 hours following standard passage (unsorted) or following sorting by either FACS or MACS in two representative NPC lines with different survival responses. Scale bar = 100μm, *p<0.05, n.s = not significant. Error bars ± SEM.

Article Snippet: CD271- cells were re-pelleted and re-suspended in fresh MACS Separation Buffer, then labelled using the Indirect CD133 Microbead kit (Miltenyi Biotech, #130-091-895) for 15 minutes at 4°C.

Techniques: Immunofluorescence, Flow Cytometry, Expressing, Lactate Dehydrogenase Assay, Staining

Journal: PLoS ONE

Article Title: Reduced variability of neural progenitor cells and improved purity of neuronal cultures using magnetic activated cell sorting

doi: 10.1371/journal.pone.0213374

Figure Lengend Snippet: (A-B). Clustering of individual cell lines using multidimensional scaling analysis based on gene expression profiles from custom designed Taqman microfluidic cards. (A)Both FACS (red) and MACS (blue) increase gene expression homogeneity between NPC lines (indicated by black circle) compared to unsorted cells (green). (B) MACS (triangles) also improves gene expression homogeneity across different neuron populations (red), as indicated by the red oval, compared to unsorted cells (squares). NPC populations are colored in blue and the homogeneous cluster is indicated by a blue circle, FACS NPCs are circles. Neurons differentiated from the unsorted F12444-3-2 NPC line that failed both FACS and MACS sorting due to low levels of CD271-/CD133+ cells are indicated with a black arrow. (C) Comparison of gene expression of glial and neuronal specific markers in neurons derived from MACS NPCs compared to neurons derived from unsorted NPCs. (D) Representative immunofluorescence images of neurons derived from NPCs demonstrating the enrichment of neuronal markers Tuj1, NeuN and Tau, as well as the depletion of glial markers GFAP and S100β when differentiated from MACS NPCs compared to unsorted NPCs. Cell nuclei are labelled with DAPI. Scale bar = 100μm, N = 5. (E-F) Flow cytometry analysis for neuronal marker TUJ1 (E) and NeuN (F) on neurons derived from MACS NPCs compared to neurons from unsorted NPCs. Relative index for each marker is generated by the multiplication of total number of fluorescent positive cells and its median fluorescence intensity. (G) Relative index of astrocyte marker, S100β by flow cytometry analysis. *p<0.05, **p<0.01, n.s = not significant. Error bars ± SEM.

Article Snippet: CD271- cells were re-pelleted and re-suspended in fresh MACS Separation Buffer, then labelled using the Indirect CD133 Microbead kit (Miltenyi Biotech, #130-091-895) for 15 minutes at 4°C.

Techniques: Gene Expression, Comparison, Derivative Assay, Immunofluorescence, Flow Cytometry, Marker, Generated, Fluorescence

Journal: PLoS ONE

Article Title: Reduced variability of neural progenitor cells and improved purity of neuronal cultures using magnetic activated cell sorting

doi: 10.1371/journal.pone.0213374

Figure Lengend Snippet: (A) Representative immunofluorescence images from a late passage NPC line demonstrating a visible reduction in CD271+ cells and an enrichment of CD133+ positive cells, as well as an enrichment of NPC markers following MACS. Cell nuclei are labelled with DAPI. Scale bar = 100μm. (B-C) Percentage of late passage NPCs positive for SOX2 (B) and NESTIN (C) as measured by flow cytometry analysis. The summary result is shown in unsorted and MACS NPCs compared to the secondary antibody control and CD271+ cells by pooled results of individual cell lines from . (D) Percent NESTIN+ cells by flow cytometry analysis in early (p2-3) and late passage (p10-13) NPCs. (E) Clustering of individual late passage NPC lines using multidimensional scaling analysis based on gene expression profiles from custom designed Taqman microfluidic cards. MACS (triangles) improves homogeneity across late passage (blue) cell lines compared to unsorted cell lines (squares), and exhibits similar gene expression profiles as early passage (red) MACS and FACS (circles) NPCs, indicated by a dotted circle. (F) Representative immunofluorescence images of neurons differentiated from one late passage NPC line demonstrating an enrichment of neuronal specific markers following MACS, but a maintenance of expression of glial-specific markers. Cell nuclei are labelled with DAPI. Scale bar = 100μm. (G) Comparison of gene expression of glial and neuronal specific markers in neurons derived from late passage MACS NPCs compared to neurons derived from early passage NPCs. *p<0.05, **p<0.01, ***p<0.001. Error bars ± SEM.

Article Snippet: CD271- cells were re-pelleted and re-suspended in fresh MACS Separation Buffer, then labelled using the Indirect CD133 Microbead kit (Miltenyi Biotech, #130-091-895) for 15 minutes at 4°C.

Techniques: Immunofluorescence, Flow Cytometry, Control, Gene Expression, Expressing, Comparison, Derivative Assay

Journal: bioRxiv

Article Title: Synthesis, anticancer properties, and biological profiling of synthetic glycan analogs of proscillaridin A

doi: 10.1101/2025.03.31.646140

Figure Lengend Snippet: Flow cytometry profiling of key markers and cell cycle analysis in human and murine colon cancer cells. A) HCT-116 cells treated with 1 μM of compound 5 exhibit significantly enhanced surface expression of cleaved caspase-3, the hallmark of pre-apoptotic cell death. B) Effect of compounds 1 through 5 on surface CD133 (prominin 1) expression. C) Effect of compounds 1 through 5 on surface CD117 (c-Kit) expression. D) Cell cycle analysis by flow cytometry on HCT-116, HT-29, and CT26 cells treated with 100 nM of compounds 1 through 5

Article Snippet: Cleaved caspase-3 anti-mouse monoclonal antibody (Cat. #Ab156544) and CD117/c-Kit anti-mouse monoclonal antibody (Cat. #Ab096639) were obtained from Aladdin Scientific Corp. Anti-CD133 monoclonal antibody HB#7 was obtained from the DSHB and deposited there by Swaminathan, S.K./Panyam, J./Ohlfest, J.R. Ribonuclease A was obtained from Research Products International (97% purity).

Techniques: Flow Cytometry, Cell Cycle Assay, Expressing