Journal: Molecular cancer

Article Title: Identification of anti-tumour biologics using primary tumour models, 3-D phenotypic screening and image-based multi-parametric profiling.

doi: 10.1186/s12943-015-0415-0

Figure Lengend Snippet: Fig. 1 Phenotypic screening in primary NSCLC tumour cells. a Images of cells derived from NSCLC primary tumour #1 cultured in three different conditions. b Effects of the scFv-Fc antibody panel upon NSCLC tumour #1 cell growth in three culture conditions, measured by Cell-Titre Glo (CTG) luminescence signal. Each antibody was individually dosed (without normalising concentrations) and cells were grown in 96-well plates. Positive (anti-IGF1R) and negative control antibodies were dosed in multiple replicates on each plate to establish consistency between plates. Each data point indicates a single well. Black horizontal bars indicate the average value for a sample class. c Scatter plot comparing the effects of scFv-Fc antibodies on NSCLC tumour #1 cell growth grown in spheroids and in standard monolayer cultures. Each data point indicates a single antibody (or replicate of the controls). The dashed box indicates a group of antibodies that strongly inhibited growth of cell monolayers but not spheroids. The solid-line box indicates a group of antibodies with a weak inhibitory effect in both spheroids and monolayers. The orange-coloured datapoint represents an antibody that was later shown to bind CDCP1 (αCDCP1-Ab3 in Fig. 2)

Article Snippet: After blocking, the NCI-H358 cells were stained for 30 min on ice with either DARPin-Fcs at 1 μg/106 cells; allophycocyanin conjugated anti-CDCP1 antibody (R&D Systems FAB26662A/Lot LVQ0109021X) at 10 μL/106 cells, or an isotype control antibody at 1 μg/106 cells.

Techniques: Derivative Assay, Cell Culture, Negative Control

Journal: bioRxiv

Article Title: Modeling signaling-dependent pluripotent cell states with boolean logic can predict cell fate transitions

doi: 10.1101/115683

Figure Lengend Snippet: (a) Predicted population-level gene expression levels (left panel) and qRT-PCR-measured relative expressions (right panel; shown in fold-change from the levels of 2iL) of OSN and lineage markers. Data represents the mean and s.e.m. of three or four biological replicates; the differences between 2iJ and 2iJ+B were examined using a 2-tailed unpaired Student’s t test and asterisks indicate *p < 0.1 and ** p <0.05. (b) In silico subpopulation analysis via threshold-based characterization for individual SCCs under the input condition of 2i-L+B-A. Stable grouped profiles enriched as either PSC, TE, ME, PE or Epiblast-like subpopulations were traced in color-coded circles. The circles with solid line indicate SCCs with no outgoing edges (sustainability score =1.0), and those with dashed line indicate SCCs with lower sustainability. (c) Confocal images of immuno-staining of mESCs for Oct4/Cdx2/Gata4 (top) or Oct4/Cdx2/Brachyury (T) (bottom panels) cultured in the given conditions for 2 days. (d) Quantification of frequency of subpopulations that exhibit features of differentiation lineages. Data are means of three replicates and the results were confirmed in two independent studies. (e) qRT-PCR for pluripotency and extended TE-lineage maker genes. TSCs and mESCs after culture in each condition were compared. Data represents the mean of three replicates. (f) Flow cytometry histograms showing fluorescence intensity of CDCP1 and CD40 in individual samples of mESC in LS, TS, and mESCs cultured in 2iJ+B-A for 2 days. Percentage listed is that of positive cells in the 2iJ+B-A condition. (g) PCA plot of RNA-seq data for the top 40% of genes that show highest variance across all samples. Distinct cell types and conditions are indicated with different colors. Circles include day2 and day5 samples for 2i-L and 2iJ conditions, and two day 2 samples, day 5 and 11 for 2iJ+B-A condition. Diamonds indicate stable cell type no culture time defined. Meso indicates mesoderm progenitors. (h) Comparison of predicted gene expression levels and RNAseq-measured gene counts in 2iJ+B-A relative to 2iJ (Equivalent to 2i-L in simulation) for 29 genes involved in the model. The experimental mean relative gene expressions of day2 samples for the two conditions and the mean relative predicted levels are shown. Black dots indicate genes significantly up- or down-regulated (p < 0.05) in three 2iJ+B-A-treated samples compared with two 2iJ-treated samples. Genes in blue are up-regulated in TSC, and those in red are down-regulated in TSC compared with 2iJ-treated samples. (i) Left panel: In vivo lineage contribution frequency and chimera efficiency of H2B-GFP ESCs treated with either 2iL or 2iJ in the presence of serum. Lineage contribution efficiencies were calculated as number of chimeras with cells in Epiblast (EPI) or TE positions/total number of chimeras. Note that cells scored as “TE-position” did not express TE marker Cdx2. Chimera forming efficiency was calculated as number of chimeras/number of total aggregates made. Right panel: Representative images of aggregation chimeras at E4.5. We observed a number of cells in TE positions in chimeras. These cells ranged from live-looking to apoptotic, however none expressed Cdx2 and thus were not considered as viable, integrated contributions to the TE lineage.

Article Snippet: Cells (mESCs and TSCs) were first stained for surface markers CDCP1 (R&D Systems AF4515) and CD40 (BD Biosciences 562846) using antibodies at 1:100 dilutions and assayed using flow cytometry (BD LSRFortessa).

Techniques: Gene Expression, Quantitative RT-PCR, In Silico, Immunostaining, Cell Culture, Flow Cytometry, Fluorescence, RNA Sequencing, Stable Transfection, Comparison, In Vivo, Marker

Journal: bioRxiv

Article Title: Modeling signaling-dependent pluripotent cell states with boolean logic can predict cell fate transitions

doi: 10.1101/115683

Figure Lengend Snippet: (a) In silico subpopulation analysis of possible signaling input combinations with 2iL and 2i-L. The threshold values for predicted expression levels of lineage specifiers in each SCC are set as follows: Oct4=0.3, EpiTFs=0.2, and Gata6=0.5 and Cdx2=0.7. (b) Frequency of Cdx2+ population including TE-like sub-population (Cdx2+/Oct4-) after five days in culture measured by immuno-staining. Data represents the means of six replicates consisting of two biological replicates with three technical replicates in each. The representative density plot of immuno-staining of Cdx2 on day 5 is shown in the left panel. (c) Histograms for expression of TE-enriched cell surface markers, CD40 and CDCP1. Shown are mESCs treated in 2iJ (1 st column), 2i-L (2 nd column), or 2iL(3 rd column), in unsupplemented medium (1 st row) or BMP4 and ALKi supplemented medium (+B-A) (2 nd row). Control mESCs (kept in control LS) and TSCs are shown at the bottom. (d) In vivo lineage contribution frequency and chimera efficiency of H2B-Tomato ESCs treated with either 2iL or 2iJ in the presence of serum.

Article Snippet: Cells (mESCs and TSCs) were first stained for surface markers CDCP1 (R&D Systems AF4515) and CD40 (BD Biosciences 562846) using antibodies at 1:100 dilutions and assayed using flow cytometry (BD LSRFortessa).

Techniques: In Silico, Expressing, Immunostaining, Control, In Vivo

Journal: bioRxiv

Article Title: Modeling Post-Gastrula Development via Bidirectional Pluripotent Stem Cells

doi: 10.1101/2025.06.28.662107

Figure Lengend Snippet: (A) Schematic of BPSC (derived from EPSC), ESC, and EPSC differentiated spontaneously without inducing factors for 2 days. (B) Typical brightfield images of BPSC (derived from EPSC), ESC, and EPSC after two days of spontaneous differentiation without inducing factors. (C) Immunofluorescence staining of BPSC (derived from EPSC), ESC, and EPSC differentiated spontaneously for 2 days. Staining for OCT4, CDX2 and GATA6. Scale bar, 50 μm. (D) Representative FACS analysis of the BPSC (derived from EPSC), ESC, and EPSC differentiated spontaneously for 2 days. (E) UMAP plot of scRNA-seq data from E4.5 embryo cells, BPSC (derived from EPSC) after 1 or 2 days of spontaneous differentiation, and EPSC after 2 days of spontaneous differentiation. Cells were colored according their cell types and split by their origins. (F) Schematic of BPSC differentiation induced with TSM for 2-3 days. After 3 days of induction, CDX2-mCherry-positive or CDCP1-positive cells were collected by FACS and cultured with TSM. After 2 days of induction, chimeras were performed with cells injected into embryos at 8-cell stage. (G) Immunofluorescence staining of BPSC-TSC (derived from EPSC), BPSC-TSC (derived from ESC) and TSCs (derived from embryos). Staining for SOX2 and TFAP2C. Scale bar, 20 μm. (H) Immunofluorescence staining of E6.5 chimeric embryo produced using the above method . Staining for TFAP2C and GFP. Scale bar, 50 μm. BPSC and ESC are labeled by GFP. Enlarged view of the blue dotted box (right panel). (I) Immunofluorescence staining of chimeric placentas at E12.5 stage. Staining for TPBPA and GFP. Scale bar, 500 μm.

Article Snippet: For the induced differentiation of BPSC BPSC differentiation with TSM with 25μg/mL FGF4 and 1μg/mL Heparin for 3 days and collected CDX2-mCherry positive or CDCP1 (R&D Systems AF4515-SP) positive cells with FACS.

Techniques: Derivative Assay, Immunofluorescence, Staining, Cell Culture, Injection, Produced, Labeling

Journal: bioRxiv

Article Title: Modeling Post-Gastrula Development via Bidirectional Pluripotent Stem Cells

doi: 10.1101/2025.06.28.662107

Figure Lengend Snippet: (A) Representative FACS analysis of the percentages of CDCP1 positive cells in EPSC, ESC, TSC, BPSC(EPSC)-TSMD3 (EPSC-derived BPSC induced with TSM for 3 days) and BPSC(ESC)-TSMD3 (ESC-derived BPSCs induced with TSM for 3 days) (B) Immunofluorescence staining of BPSC-TSC (EPSC), BPSC-TSC (ESC) and TSC (derived from embryo). Staining for SOX2 and CDX2. Scale bar, 20 μm. (C) Representative morphological images of E6.5 chimeric embryo produced using the above method . (D) E6.5 Chimera embryo formation ratio of differentiated BPSC induced with TSM for 2 days and ESC. (E) Representative image of ESC and BPSC-diff-D2 chimera at E12.5 stage.

Article Snippet: For the induced differentiation of BPSC BPSC differentiation with TSM with 25μg/mL FGF4 and 1μg/mL Heparin for 3 days and collected CDX2-mCherry positive or CDCP1 (R&D Systems AF4515-SP) positive cells with FACS.

Techniques: Derivative Assay, Immunofluorescence, Staining, Produced

Journal: International Journal of Biological Sciences

Article Title: Osimertinib and pterostilbene in EGFR-mutation-positive non-small cell lung cancer (NSCLC)

doi: 10.7150/ijbs.32889

Figure Lengend Snippet: Used antibodies, including dilutions and catalog numbers.

Article Snippet: Rabbit anti-Phospho-CDCP1 (Y707) , 1:1000 , Cell Signaling (#13111).

Techniques: