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ATCC mcf10a cells
Mcf10a Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 7956 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mcf10a+cells/us12629375-542-3-8?v=ATCC
Average 99 stars, based on 7956 article reviews

mcf10a cells - by Bioz Stars, 2026-07

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a.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells. Colors shown in the key and dotted lines denote gating used for analysis throughout this publication. b.) Scatterplot of DNA content versus median cyclin B1 intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Colors shown in the key and dotted lines denote cyclin B1 gating used for analysis throughout this article. c.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Colors shown in the key and dotted lines denote cyclin B1 gating used for analysis throughout this article. d.) Quantification of mean cyclin B1 intensity in MCF10A cells with non-targeting or cyclin B1 knockdowns to demonstrate cyclin B1 antibody specificity. A Mann-Whitney test was conducted to assess significance. Error bars are representative of 10-90 percentile range. P value: <0.0001. e.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle from RPE-1 cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. f.) Quantification of integrated SLBP intensity in MCF10A cells with non-targeting or SLBP knockdowns to demonstrate SLBP antibody specificity. Error bars are representative of 10-90 percentile range. A Mann-Whitney test was conducted to assess significance. P value: <0.0001. g.) Breakdown of quantification of median SLBP nuclear intensity across the cell cycle from into individual graphs. Data are presented as mean ± SEM. h.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, 5μM RO-3306, or a combination of 5μM AZD6738 and 5μM RO-3306 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM. i.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, 1μM NU6140, or a combination of 5μM AZD6738 and 1μM NU6140 for 1hrs (n=3 biological replicates). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells. Colors shown in the key and dotted lines denote gating used for analysis throughout this publication. b.) Scatterplot of DNA content versus median cyclin B1 intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Colors shown in the key and dotted lines denote cyclin B1 gating used for analysis throughout this article. c.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Colors shown in the key and dotted lines denote cyclin B1 gating used for analysis throughout this article. d.) Quantification of mean cyclin B1 intensity in MCF10A cells with non-targeting or cyclin B1 knockdowns to demonstrate cyclin B1 antibody specificity. A Mann-Whitney test was conducted to assess significance. Error bars are representative of 10-90 percentile range. P value: <0.0001. e.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle from RPE-1 cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. f.) Quantification of integrated SLBP intensity in MCF10A cells with non-targeting or SLBP knockdowns to demonstrate SLBP antibody specificity. Error bars are representative of 10-90 percentile range. A Mann-Whitney test was conducted to assess significance. P value: <0.0001. g.) Breakdown of quantification of median SLBP nuclear intensity across the cell cycle from into individual graphs. Data are presented as mean ± SEM. h.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, 5μM RO-3306, or a combination of 5μM AZD6738 and 5μM RO-3306 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM. i.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, 1μM NU6140, or a combination of 5μM AZD6738 and 1μM NU6140 for 1hrs (n=3 biological replicates). Data are presented as mean ± SEM.

Article Snippet: Human MCF10A cells (ATCC, CRL-10317) were grown in DMEM/F12 medium (ThermoFisher Scientific, 11330057) supplemented with 100U/mL penicillin/streptomycin (Thermo Fisher Scientific, 15140122), 5% horse serum (ThermoFisher Scientific, 16050122), 20ng/ml epidermal growth factor (EGF) (Sigma-Aldrich, E5036), 0.5 mg/mL hydrocortisone, 10μg/mL insulin (Sigma-Aldrich, I1882-100MG), and 100ng/mL cholera toxin (Sigma-Aldrich, C8052).

Techniques: MANN-WHITNEY

a.) Representative images of cyclin B1 and EdU in MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Arrows indicate nuclei positive for both EdU and cyclin B1. Scale bar is equal to 40μm. b.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Data are presented as mean ± SEM. c.) Scatterplots of DNA content (integrated DAPI intensity) versus mean cyclin B1 cytoplasmic intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by EdU intensity (gating of negative/positive EdU values in ) as denoted by the associated scale. Representative of n=4 biological replicates. d.) Scatterplots of DNA content (integrated DAPI intensity) versus mean EdU intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by mean cyclin B1 intensity as denoted by the associated scale. Representative of n=4 biological replicates. e.) Quantification of mean cyclin B1 intensity values from in EdU positive cells. Error bars are representative of 10-90 percentile range. A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: <0.0001 (DMSO versus AZD6738) and <0.0001 (DMSO versus LY2603618). f.) Quantification of mean EdU intensity values from in cells with high cyclin B1 levels (gating of low/high cyclin B1 values in ). Error bars are representative of 10-90 percentile range. P values: <0.0001 (DMSO versus AZD6738) and <0.0001 (DMSO versus LY2603618). g.) Representative images of SLBP and EdU in S phase MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=3 biological replicates. Scale bar is equal to 10μm. h.) Quantification of median SLBP nuclear intensity across the cell cycle in MCF10A cells treated with DMSO, 5μM AZD6738, 5μM VE822, 2μM LY2603618, or 250nM ChIR124 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Representative images of cyclin B1 and EdU in MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Arrows indicate nuclei positive for both EdU and cyclin B1. Scale bar is equal to 40μm. b.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Data are presented as mean ± SEM. c.) Scatterplots of DNA content (integrated DAPI intensity) versus mean cyclin B1 cytoplasmic intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by EdU intensity (gating of negative/positive EdU values in ) as denoted by the associated scale. Representative of n=4 biological replicates. d.) Scatterplots of DNA content (integrated DAPI intensity) versus mean EdU intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by mean cyclin B1 intensity as denoted by the associated scale. Representative of n=4 biological replicates. e.) Quantification of mean cyclin B1 intensity values from in EdU positive cells. Error bars are representative of 10-90 percentile range. A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: <0.0001 (DMSO versus AZD6738) and <0.0001 (DMSO versus LY2603618). f.) Quantification of mean EdU intensity values from in cells with high cyclin B1 levels (gating of low/high cyclin B1 values in ). Error bars are representative of 10-90 percentile range. P values: <0.0001 (DMSO versus AZD6738) and <0.0001 (DMSO versus LY2603618). g.) Representative images of SLBP and EdU in S phase MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=3 biological replicates. Scale bar is equal to 10μm. h.) Quantification of median SLBP nuclear intensity across the cell cycle in MCF10A cells treated with DMSO, 5μM AZD6738, 5μM VE822, 2μM LY2603618, or 250nM ChIR124 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM.

Techniques:

a.) Representative images of cyclin B1, EdU, and yH2AX in MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Arrows indicate nuclei positive for EdU, cyclin B1 and yH2AX. Scale bar is equal to 50μm. b.) Scatterplots of DNA content (integrated DAPI intensity) versus mean yH2AX intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by high or low cyclin B1 intensity as denoted by the associated scale. Representative of n=4 biological replicates. c.) Quantification of median cyclin B1 cytoplasmic intensity in MCF10A cells with low or high yH2AX across the cell cycle. Cells were treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of mitotic failure (defined in ) in control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Percentages were calculated based on counts of 4 images per treatment for each biological replicate. A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.9964 (siControl versus siCCNB1 + DMSO), <0.0001 (siControl versus siCCNB1 + AZD6738), and 0.0003 (siControl versus siCCNB1 + LY2603618). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Representative images of cyclin B1, EdU, and yH2AX in MCF10A cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Arrows indicate nuclei positive for EdU, cyclin B1 and yH2AX. Scale bar is equal to 50μm. b.) Scatterplots of DNA content (integrated DAPI intensity) versus mean yH2AX intensity in MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Each dot is representative of values from a singular cell. The plot is colored by high or low cyclin B1 intensity as denoted by the associated scale. Representative of n=4 biological replicates. c.) Quantification of median cyclin B1 cytoplasmic intensity in MCF10A cells with low or high yH2AX across the cell cycle. Cells were treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of mitotic failure (defined in ) in control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Percentages were calculated based on counts of 4 images per treatment for each biological replicate. A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.9964 (siControl versus siCCNB1 + DMSO), <0.0001 (siControl versus siCCNB1 + AZD6738), and 0.0003 (siControl versus siCCNB1 + LY2603618). Data are presented as mean ± SEM.

Techniques: Control, Knockdown

a.) Quantification of median cyclin B1 cytoplasmic intensity in RPE-1 cells with low or high yH2AX across the cell cycle. Cells were treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. b.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control and cyclin B1 knockdown RPE-1 cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. c.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM KU60019 + 5μM AZD6738, 5μM KU60019 + or 2μM LY2603618 + 5μM KU60019 for 7hrs (n=3 biological replicates). Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM NU7441 + 5μM AZD6738, 5μM NU7441 + or 2μM LY2603618 + 5μM NU7441 for 7hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Representative images of cells undergoing normal mitosis versus mitotic failure. Scale bar is equal to 15μm.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Quantification of median cyclin B1 cytoplasmic intensity in RPE-1 cells with low or high yH2AX across the cell cycle. Cells were treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. b.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control and cyclin B1 knockdown RPE-1 cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. c.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM KU60019 + 5μM AZD6738, 5μM KU60019 + or 2μM LY2603618 + 5μM KU60019 for 7hrs (n=3 biological replicates). Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM NU7441 + 5μM AZD6738, 5μM NU7441 + or 2μM LY2603618 + 5μM NU7441 for 7hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Representative images of cells undergoing normal mitosis versus mitotic failure. Scale bar is equal to 15μm.

Techniques: Control, Knockdown

a.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells. Colors shown in the key and dotted lines denote gating of EdU negative cells with >3n DNA content (referred to as G2) used for analysis of chromatin-bound replication component retention. b.) Scatterplots of mean chromatin-bound PCNA intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The dotted line and color of the points indicate the gating for PCNA positive and negative cells. c.) Scatterplots of mean EdU intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The points are colored by status of being PCNA negative/positive as denoted by the associated key. Representative of n=3 biological replicates. d.) Scatterplots of mean chromatin-bound MCM2 intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The dotted line and color of the points indicate the gating for MCM2 positive and negative cells. E.) Scatterplots of mean EdU intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The points are colored by status of being MCM2 negative/positive as denoted by the associated key. Representative of n=3 biological replicates.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Scatterplot of DNA content versus mean EdU intensity in MCF10A cells. Colors shown in the key and dotted lines denote gating of EdU negative cells with >3n DNA content (referred to as G2) used for analysis of chromatin-bound replication component retention. b.) Scatterplots of mean chromatin-bound PCNA intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The dotted line and color of the points indicate the gating for PCNA positive and negative cells. c.) Scatterplots of mean EdU intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The points are colored by status of being PCNA negative/positive as denoted by the associated key. Representative of n=3 biological replicates. d.) Scatterplots of mean chromatin-bound MCM2 intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The dotted line and color of the points indicate the gating for MCM2 positive and negative cells. E.) Scatterplots of mean EdU intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The points are colored by status of being MCM2 negative/positive as denoted by the associated key. Representative of n=3 biological replicates.

Techniques:

a.) Scatterplots of mean chromatin-bound PCNA intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. b.) Quantification of mean PCNA intensities from in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population (Supplemental Fig. 3a). Error bars are representative of 10-90 percentile range. c.) Scatterplots of mean MCM2 intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. d.) Quantification of mean MCM2 intensities from in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population. Error bars are representative of 10-90 percentile range. e.) Quantification of integrated DAPI intensities to measure DNA content in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population (n=3 biological replicates). Error bars are representative of 10-90 percentile range. f.) Quantification of the percentage of PCNA (left) or MCM2 (right) positive pre-extracted MCF10A cells (gating of PCNA and MCM2 shown in and ) in the defined G2 population of control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed for each plot to assess biological significance. P values for PCNA plot: >0.9999 (DMSO siControl versus siCCNB1), <0.0001 (AZD6738 siControl versus siCCNB1), and <0.0001 (LY2603618 siControl versus siCCNB1). P values for MCM2 plot: 0.9694 (DMSO siControl versus siCCNB1), 0.0021 (AZD6738 siControl versus siCCNB1), and 0.0081 (LY2603618 siControl versus siCCNB1). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Scatterplots of mean chromatin-bound PCNA intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. b.) Quantification of mean PCNA intensities from in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population (Supplemental Fig. 3a). Error bars are representative of 10-90 percentile range. c.) Scatterplots of mean MCM2 intensity versus DNA content in pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. d.) Quantification of mean MCM2 intensities from in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population. Error bars are representative of 10-90 percentile range. e.) Quantification of integrated DAPI intensities to measure DNA content in pre-extracted MCF10A cells with low or high yH2AX in the defined G2 population (n=3 biological replicates). Error bars are representative of 10-90 percentile range. f.) Quantification of the percentage of PCNA (left) or MCM2 (right) positive pre-extracted MCF10A cells (gating of PCNA and MCM2 shown in and ) in the defined G2 population of control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed for each plot to assess biological significance. P values for PCNA plot: >0.9999 (DMSO siControl versus siCCNB1), <0.0001 (AZD6738 siControl versus siCCNB1), and <0.0001 (LY2603618 siControl versus siCCNB1). P values for MCM2 plot: 0.9694 (DMSO siControl versus siCCNB1), 0.0021 (AZD6738 siControl versus siCCNB1), and 0.0081 (LY2603618 siControl versus siCCNB1). Data are presented as mean ± SEM.

Techniques: Control, Knockdown

a.) Representative images of PCNA, yH2AX, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Cell cycle status is indicated in the figure. Representative of n=3 biological replicates. Scale bar is equal to 10μm. b.) Representative images of MCM2, yH2AX, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Cell cycle status is indicated in the figure. Representative of n=3 biological replicates. Scale bar is equal to 10μm. C.) Representative images of PCNA, MCM2, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The DMSO treated nuclei shows a cell in late S phase, while the nuclei from the other treatments show EdU negative cells that have halted replication. Whit boxes indicate areas of the nuclei that have been magnified. Representative of n=3 biological replicates. Scale bar is equal to 10μm.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Representative images of PCNA, yH2AX, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Cell cycle status is indicated in the figure. Representative of n=3 biological replicates. Scale bar is equal to 10μm. b.) Representative images of MCM2, yH2AX, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Cell cycle status is indicated in the figure. Representative of n=3 biological replicates. Scale bar is equal to 10μm. C.) Representative images of PCNA, MCM2, and EdU in pre-extracted MCF10A nuclei treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. The DMSO treated nuclei shows a cell in late S phase, while the nuclei from the other treatments show EdU negative cells that have halted replication. Whit boxes indicate areas of the nuclei that have been magnified. Representative of n=3 biological replicates. Scale bar is equal to 10μm.

Techniques:

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Techniques:

a.) Quantification the percentage of PCNA positive pre-extracted cells in the defined G2 population of control, USP37, or TRAIP knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.5397 (DMSO siControl versus siUSP37), 0.6511 (DMSO siControl versus siTRAIP), <0.0001 (AZD6738 siControl versus siUSP37), 0.0068 (AZD6738 siControl versus siTRAIP), 0.0003 (LY2603618 siControl versus siUSP37), and 0.0101 (LY2603618 siControl versus siUSP37). Data are presented as mean ± SEM. b.) Scatterplots of mean PCNA intensity versus DNA content in control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. c.) Quantification of the percentage of MCM2 positive pre-extracted cells in the defined G2 population of control, USP37, or TRAIP knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.0837 (DMSO siControl versus siUSP37), 0.8341 (DMSO siControl versus siTRAIP), <0.0001 (AZD6738 siControl versus siUSP37), 0.0433 (AZD6738 siControl versus siTRAIP), 0.0410 (LY2603618 siControl versus siUSP37), and 0.5357 (LY2603618 siControl versus siUSP37). Data are presented as mean ± SEM. d.) Scatterplots of mean MCM2 intensity versus DNA content in control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. e.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Quantification the percentage of PCNA positive pre-extracted cells in the defined G2 population of control, USP37, or TRAIP knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.5397 (DMSO siControl versus siUSP37), 0.6511 (DMSO siControl versus siTRAIP), <0.0001 (AZD6738 siControl versus siUSP37), 0.0068 (AZD6738 siControl versus siTRAIP), 0.0003 (LY2603618 siControl versus siUSP37), and 0.0101 (LY2603618 siControl versus siUSP37). Data are presented as mean ± SEM. b.) Scatterplots of mean PCNA intensity versus DNA content in control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. c.) Quantification of the percentage of MCM2 positive pre-extracted cells in the defined G2 population of control, USP37, or TRAIP knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.0837 (DMSO siControl versus siUSP37), 0.8341 (DMSO siControl versus siTRAIP), <0.0001 (AZD6738 siControl versus siUSP37), 0.0433 (AZD6738 siControl versus siTRAIP), 0.0410 (LY2603618 siControl versus siUSP37), and 0.5357 (LY2603618 siControl versus siUSP37). Data are presented as mean ± SEM. d.) Scatterplots of mean MCM2 intensity versus DNA content in control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Plots are divided by EdU negative/positive cells and by treatment. The plot is colored by mean yH2AX intensity as denoted by the associated scale. Representative of n=3 biological replicates. e.) Quantification of mean yH2AX nuclear intensities across the cell cycle from control, USP37, or TRAIP knockdown and pre-extracted MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM.

Techniques: Control, Knockdown

a.) Schematic of ATRi resistant cell line development (ATRi-R). b.) Cell proliferation plot of ATRi-R cells compared to controls under 5μM AZD6738 treatment. c.) Representative images of yH2AX in MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Representative of n=3 biological replicates. Scale bar is equal to 50μm. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 1mM hydroxyurea or a combination of 5μM AZD6738 and 1mM hydroxyurea 1.5hrs (n=3 biological replicates). Data are presented as mean ± SEM. f.) Representative images of DAPI stained nuclei in MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Representative of n=3 biological replicates. Arrows indicate cells with mitotic slippage. Scale bar is equal to 20μm. g.) Quantification of mitotic failure (defined in ) in control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.9983 (DMSO MCF10A versus ATRi-R), 0.0027 (AZD6738 MCF10A versus ATRi-R), 0.0007 (LY2603618 MCF10A versus ATRi-R). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Schematic of ATRi resistant cell line development (ATRi-R). b.) Cell proliferation plot of ATRi-R cells compared to controls under 5μM AZD6738 treatment. c.) Representative images of yH2AX in MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Representative of n=3 biological replicates. Scale bar is equal to 50μm. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of mean yH2AX nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 1mM hydroxyurea or a combination of 5μM AZD6738 and 1mM hydroxyurea 1.5hrs (n=3 biological replicates). Data are presented as mean ± SEM. f.) Representative images of DAPI stained nuclei in MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs. Representative of n=3 biological replicates. Arrows indicate cells with mitotic slippage. Scale bar is equal to 20μm. g.) Quantification of mitotic failure (defined in ) in control and cyclin B1 knockdown MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=3 biological replicates). A two-way analysis of variance (ANOVA) was performed to assess biological significance. P values: 0.9983 (DMSO MCF10A versus ATRi-R), 0.0027 (AZD6738 MCF10A versus ATRi-R), 0.0007 (LY2603618 MCF10A versus ATRi-R). Data are presented as mean ± SEM.

Techniques: Staining, Control, Knockdown

a.) Quantification of mean yH2AX nuclear intensity across the cell cycle from ATRi-R cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM KU60019 + 5μM AZD6738, 5μM KU60019 + or 2μM LY2603618 + 5μM KU60019 for 7hrs (n=3 biological replicates). Data from the parental MCF10A cell line is also presented in Supplementary Fig. 1c. Arrows indicate plots of ATRi-R values with scaled Y axis. Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from ATRi-R cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM NU7441 + 5μM AZD6738, 5μM NU7441 + or 2μM LY2603618 + 5μM NU7441 for 7hrs (n=3 biological replicates). Data from the parental MCF10A cell line is also presented in Supplementary Fig. 1d. Arrows indicate plots of ATRi-R values with scaled Y axis. Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Quantification of mean yH2AX nuclear intensity across the cell cycle from ATRi-R cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM KU60019 + 5μM AZD6738, 5μM KU60019 + or 2μM LY2603618 + 5μM KU60019 for 7hrs (n=3 biological replicates). Data from the parental MCF10A cell line is also presented in Supplementary Fig. 1c. Arrows indicate plots of ATRi-R values with scaled Y axis. Data are presented as mean ± SEM. d.) Quantification of mean yH2AX nuclear intensity across the cell cycle from ATRi-R cells treated with DMSO + DMSO, DMSO + 5μM AZD6738, DMSO + 2μM LY2603618, 5μM KU60019 + DMSO, 5μM NU7441 + 5μM AZD6738, 5μM NU7441 + or 2μM LY2603618 + 5μM NU7441 for 7hrs (n=3 biological replicates). Data from the parental MCF10A cell line is also presented in Supplementary Fig. 1d. Arrows indicate plots of ATRi-R values with scaled Y axis. Data are presented as mean ± SEM.

Techniques:

a.) Representative images of cyclin B1, EdU, and yH2AX in MCF10A and ATRi-R cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Scale bar is equal to 50μm. b.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Note: MCF10A data was also shown in Fig.1b. Data are presented as mean ± SEM. c.) Representative images of pFOXM1 (T600) and EdU in MCF10A and ATRi-R cells treated with DMSO or 5μM AZD6738 for 1hr. Representative of n=3 biological replicates. Scale bar is equal to 10μm. d.) Quantification of median pFOXM1 (T600) nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM.

Journal: bioRxiv

Article Title: ATR enforcement of the S/G2 checkpoint prevents premature S phase shutdown and genome instability

doi: 10.64898/2026.05.07.723638

Figure Lengend Snippet: a.) Representative images of cyclin B1, EdU, and yH2AX in MCF10A and ATRi-R cells treated with DMSO or 5μM AZD6738 for 16hrs. Representative of n=4 biological replicates. Scale bar is equal to 50μm. b.) Quantification of median cyclin B1 cytoplasmic intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 16hrs (n=4 biological replicates). Note: MCF10A data was also shown in Fig.1b. Data are presented as mean ± SEM. c.) Representative images of pFOXM1 (T600) and EdU in MCF10A and ATRi-R cells treated with DMSO or 5μM AZD6738 for 1hr. Representative of n=3 biological replicates. Scale bar is equal to 10μm. d.) Quantification of median pFOXM1 (T600) nuclear intensity across the cell cycle from MCF10A cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM. e.) Quantification of median SLBP nuclear intensity across the cell cycle from MCF10A and ATRi-R cells treated with DMSO, 5μM AZD6738, or 2μM LY2603618 for 1hr (n=3 biological replicates). Data are presented as mean ± SEM.

Techniques:

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