# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

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# mouse monoclonal anti phospho histone h2a x ser139 (Cell Signaling Technology Inc)

Cell Signaling Technology Inc manufactures this product

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Cell Signaling Technology Inc

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

# ab4074 anti phospho histone h2a x ser139 mouse monoclonal ab (Danaher Inc)

Danaher Inc is a verified supplier

Danaher Inc manufactures this product

## Structured Review

Ab4074 Anti Phospho Histone H2a X Ser139 Mouse Monoclonal Ab, supplied by Danaher Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/ab4074 anti phospho histone h2a x ser139 mouse monoclonal ab/product/Danaher Inc

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "BRCA1 and 53BP1 Regulate Reprogramming Efficiency by Mediating DNA Repair Pathway Choice at Replication-associated Double-Strand Breaks"

**Article Title: **BRCA1 and 53BP1 Regulate Reprogramming Efficiency by Mediating DNA Repair Pathway Choice at Replication-associated Double-Strand Breaks

**Journal: **Cell reports

**doi: **10.1016/j.celrep.2024.114006

**Figure Legend Snippet:**A) Illustration of the BRCA1 polypeptide and a simplified interaction engagements. The C-terminal BRCT domain of BRCA1 interacts in a mutually exclusive manner with the phosphorylated isoforms of ABRAXAS, BACH1, or CtIP to form distinct BRCA1 complexes. In addition, BRCA1 harbors a coiled-coil motif that mediates its interaction with PALB2 and the recruitment of BRCA2 and RAD51 to sites of DNA damage. The mutant mouse alleles used in this study: Brca1tr encodes a pathogenic truncating mutation, denoted with a red arrow, that eliminates the BRCT domains. AbraxasS404A, Bach1S994A and CtipS326A each encode a missense mutation that eliminates a phosphorylation site necessary for the interaction of its protein product with the BRCT domain of BRCA1. B) Morphology and size of E13.5 embryos. The difference between wt ctrl and AABBCC was evaluated with an unpaired two-tailed student’s t-test.; wt ctrl n = 9, AABB n = 5, BBCC n = 4, AACC n = 3, AABBCC n = 3. C) Rad51 foci immunofluorescence and quantification in induced pluripotent stem (iPS) cell lines treated with 10Gy IR. Data was analyzed by one-way ANOVA.; wt ctrl n = 3, AABB n =3, BBCC n = 3, AACC n = 4, AABBCC n = 4; A = Abraxas S404A/S404A, B = Bach1 S994A/S994A, C = CtIP S326A/S326A; scale bar = 10μm. D) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cell lines, shown as a ratio of dual allele targeting in each genotype vs. control. Statistical analysis with one-way ANOVA, except for BBCC vs. AABBCC and AACC vs AABBCC, which used unpaired two-tailed student’s t-tests.; wt ctrl n = 4, AABB n = 4, BBCC n = 3, AACC n = 5, AABBCC n= 5. E) DNA fork stalling in immortalized MEFs. At least 150 DNA fibers were measured per genotype. Analysis with one-way ANOVA. F) Immunofluorescence and quantification for phospho-H2AX (S139). Foci were counted on reprogramming day 5 in > 138 cells/genotype. Statistical analysis with one-way ANOVA. Size bar = 5μm, applicable to all panels. G) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Analysis by one-way ANOVA from biological replicates. wt ctrl n = 3, AABB n = 4, BBCC n = 4, AACC n = 3, AABBCC n = 4.

**Techniques Used: **Mutagenesis, Two Tailed Test, Immunofluorescence, CRISPR, Control, Staining

**Figure Legend Snippet:**Related to Fig. S1. A) A schematic of Bard1-mediated SFP. The Bard1K607A point mutation prevents the recruitment of the Brca1/Bard1 heterodimer to reversed stalled replication forks, which makes them vulnerable to Mre11-dependent degradation. B) Immunofluorescence and quantification for double strand break (DSB) marker phospho - H2AX (S139). Foci were counted on reprogramming day 5 in ≥260 cells/genotype, statistical analysis with one-way ANOVA. scale bar = 5μm. C) Immunofluorescence and quantification of phospho -RPA(S33) on reprogramming day 5. Data from ≥240 cells/genotype and analyzed by one-way ANOVA.; The white arrows point to foci. scale bar = 5μm. D) Cell proliferation plots on reprogramming day 5. Arrested cells retain CFSE and are detectable as a bright peak by flow cytometry. Analysis by one-way ANOVA; wt ctrl n = 3, Brca1tr/+ n = 2, Bard1K607A/K607A n = 3. E) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Analysis with one-way ANOVA.; wt ctrl n = 6, Brca1tr/+ n = 6, Bard1K607A/+ n = 4, Bard1K607A/K607A n = 4, n=biological replicates.

**Techniques Used: **Mutagenesis, Immunofluorescence, Marker, Flow Cytometry, Staining

**Figure Legend Snippet:**Related to Fig. S2 A) A schematic for rescuing SFP in BRCA1 mutant cells by ablation of Smarcal1. B) DNA fiber analysis in a fork stalling assay with hydroxyurea (HU) on reprogramming day 5. At least 120 DNA fibers were measured per genotype. Analysis by one-way ANOVA. C) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cells lines. Data is shown as a ratio of dual allele targeting in each genotype relative to control. Statistical analysis with one-way ANOVA; wt ctrl n = 9, Brca1tr/+ n = 3, Brca1tr/tr n = 3, Brca1tr/tr Smarcal1−/− n = 3. D) Immunofluorescence and quantification for double strand break (DSB) marker phospho-H2AX (S139). Foci were counted on reprogramming day 5 (≥410 cells/genotype) and statistical analysis performed with one-way ANOVA. E) Immunofluorescence and quantification of ssDNA marker phospho-RPA(S33) on reprogramming day 5. Data collected from ≥140 cells per genotype and analyzed by one-way ANOVA. F) Cell proliferation analysis with CFSE on reprogramming day 5. Statistical analysis with one-way ANOVA.; wt ctrl n = 4, Smarcal1+/− n = 4, Smarcal1−/− n = 5, Brca1tr/tr n = 2, Brca1tr/tr Smarcal1−/− n = 3. G) Apoptosis analysis with Annexin V and propidium iodide (PI) on reprogramming day 5. Analysis by one-way ANOVA.; wt ctrl n = 3, Smarcal1+/− n = 3, Smarcal1−/− n = 3, Brca1tr/tr n = 3, Brca1tr/tr Smarcal1−/− n = 3. H) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Data analysis with one-way ANOVA.; wt ctrl n = 7, Brca1tr/+ n = 6, Smarcal1+/− n = 4, Smarcal1−/− n = 2, Brca1tr/tr n = 12, Brca1tr/tr Smarcal1+/− n = 4, Brca1tr/tr Smarcal1−/− n = 4. n= biological replicates.

**Techniques Used: **Mutagenesis, CRISPR, Control, Immunofluorescence, Marker, Staining

**Figure Legend Snippet:**Related to Fig. S3. A) A schematic for rescuing HDR in Brca1 mutant cells by ablation of 53bp1. Relevant substrates that can be repaired by HDR or NHEJ are shown. B) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cells lines. Data is shown as a ratio of dual allele targeting in each genotype to dual allele targeting in the control. Statistical analysis using one-way ANOVA, except for the comparison between wt ctrl and 53bp1−/−, evaluated with a two-tailed, unpaired student’s t-test.; wt ctrl n = 9, Brca1tr/+ n = 3, 53bp1−/− n = 5, Brca1tr/tr n = 3, Brca1tr/tr53bp1+/− n = 2, Brca1tr/tr53bp1−/− n = 4. C) DNA fiber analysis in a fork stalling assay with hydroxyurea (HU) on reprogramming day 5. At least 120 DNA fibers were measured per genotype and data was analyzed by one-way ANOVA. D) Immunofluorescence and quantification for phospho-H2AX (S139). Foci counted on reprogramming day 5 (≥410 cells/genotype), statistical analysis performed with one-way ANOVA. E) Immunofluorescence and quantification of ssDNA marker phospho RPA(S33) on reprogramming day 5. ≥140 cells per genotype, analyzed by one-way ANOVA. For control and BRCA1, images are identical for Fig. 3 and Fig. 4 for panels c and d. F) Cell proliferation analysis with CFSE dye on reprogramming day 5. Statistics with one-way ANOVA.; wt ctrl n = 4,, 53bp1’/- n = 5, 53bp1−/− n = 3, Brca1tr/tr n = 2, Brca1tr/trSmarcal1−/− n = 3, Brca1tr/tr 53bp1−/− n = 3. G) Apoptosis analysis with Annexin V and propidium iodide (PI) on reprogramming day 5, analyzed by one-way ANOVA.; wt ctrl n = 3, 53bp1+/− n = 3, 53bp1−/− n = 2, Brca1tr/tr n = 3, Brca1tr/tr 53bp1−/− n = 3. H) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. Number of AP positive colonies is shown as a ratio to wild type, analyzed with one-way ANOVA.; wt ctrl n = 7, Brca1tr/+ n = 6, 53bp1+/− n = 7, 53bp1−/− n = 8, Brca1tr/tr n = 12, Brca1tr/tr 53bp1−/− n = 7. I) Quantification of Nanog positive colonies in the indicated genotypes, analyzed with one-way ANOVA.; n = 3 for each genotype. J) AP staining and reprogramming efficiency quantification in human 1023 fibroblasts from adult skin biopsy in control and 53BP1 knockdown (KD) conditions. Cells were fixed on day 25 post reprogramming factor transduction, statistical analysis using an unpaired, two-tailed student’s t-test.; ctrl n = 16, 53BP1 KD n = 16. Samples with n=2 are not used for statistical comparisons. All numbers indicated are biological replicates.

**Techniques Used: **Mutagenesis, CRISPR, Control, Comparison, Two Tailed Test, Immunofluorescence, Marker, Staining, Knockdown, Transduction

**Figure Legend Snippet:**KEY RESOURCES TABLE

**Techniques Used: **Purification, Transduction, Recombinant, Knock-Out, Plasmid Preparation, Software, Imaging

# anti phospho histone h2a x ser139 mouse monoclonal ab does not react to s139a in human pluripotent stem cells (Millipore)

## Structured Review

Anti Phospho Histone H2a X Ser139 Mouse Monoclonal Ab Does Not React To S139a In Human Pluripotent Stem Cells, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/anti phospho histone h2a x ser139 mouse monoclonal ab does not react to s139a in human pluripotent stem cells/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "BRCA1 and 53BP1 Regulate Reprogramming Efficiency by Mediating DNA Repair Pathway Choice at Replication-associated Double-Strand Breaks"

**Article Title: **BRCA1 and 53BP1 Regulate Reprogramming Efficiency by Mediating DNA Repair Pathway Choice at Replication-associated Double-Strand Breaks

**Journal: **Cell reports

**doi: **10.1016/j.celrep.2024.114006

**Figure Legend Snippet:**A) Illustration of the BRCA1 polypeptide and a simplified interaction engagements. The C-terminal BRCT domain of BRCA1 interacts in a mutually exclusive manner with the phosphorylated isoforms of ABRAXAS, BACH1, or CtIP to form distinct BRCA1 complexes. In addition, BRCA1 harbors a coiled-coil motif that mediates its interaction with PALB2 and the recruitment of BRCA2 and RAD51 to sites of DNA damage. The mutant mouse alleles used in this study: Brca1tr encodes a pathogenic truncating mutation, denoted with a red arrow, that eliminates the BRCT domains. AbraxasS404A, Bach1S994A and CtipS326A each encode a missense mutation that eliminates a phosphorylation site necessary for the interaction of its protein product with the BRCT domain of BRCA1. B) Morphology and size of E13.5 embryos. The difference between wt ctrl and AABBCC was evaluated with an unpaired two-tailed student’s t-test.; wt ctrl n = 9, AABB n = 5, BBCC n = 4, AACC n = 3, AABBCC n = 3. C) Rad51 foci immunofluorescence and quantification in induced pluripotent stem (iPS) cell lines treated with 10Gy IR. Data was analyzed by one-way ANOVA.; wt ctrl n = 3, AABB n =3, BBCC n = 3, AACC n = 4, AABBCC n = 4; A = Abraxas S404A/S404A, B = Bach1 S994A/S994A, C = CtIP S326A/S326A; scale bar = 10μm. D) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cell lines, shown as a ratio of dual allele targeting in each genotype vs. control. Statistical analysis with one-way ANOVA, except for BBCC vs. AABBCC and AACC vs AABBCC, which used unpaired two-tailed student’s t-tests.; wt ctrl n = 4, AABB n = 4, BBCC n = 3, AACC n = 5, AABBCC n= 5. E) DNA fork stalling in immortalized MEFs. At least 150 DNA fibers were measured per genotype. Analysis with one-way ANOVA. F) Immunofluorescence and quantification for phospho-H2AX (S139). Foci were counted on reprogramming day 5 in > 138 cells/genotype. Statistical analysis with one-way ANOVA. Size bar = 5μm, applicable to all panels. G) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Analysis by one-way ANOVA from biological replicates. wt ctrl n = 3, AABB n = 4, BBCC n = 4, AACC n = 3, AABBCC n = 4.

**Techniques Used: **Mutagenesis, Two Tailed Test, Immunofluorescence, CRISPR, Control, Staining

**Figure Legend Snippet:**Related to Fig. S1. A) A schematic of Bard1-mediated SFP. The Bard1K607A point mutation prevents the recruitment of the Brca1/Bard1 heterodimer to reversed stalled replication forks, which makes them vulnerable to Mre11-dependent degradation. B) Immunofluorescence and quantification for double strand break (DSB) marker phospho - H2AX (S139). Foci were counted on reprogramming day 5 in ≥260 cells/genotype, statistical analysis with one-way ANOVA. scale bar = 5μm. C) Immunofluorescence and quantification of phospho -RPA(S33) on reprogramming day 5. Data from ≥240 cells/genotype and analyzed by one-way ANOVA.; The white arrows point to foci. scale bar = 5μm. D) Cell proliferation plots on reprogramming day 5. Arrested cells retain CFSE and are detectable as a bright peak by flow cytometry. Analysis by one-way ANOVA; wt ctrl n = 3, Brca1tr/+ n = 2, Bard1K607A/K607A n = 3. E) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Analysis with one-way ANOVA.; wt ctrl n = 6, Brca1tr/+ n = 6, Bard1K607A/+ n = 4, Bard1K607A/K607A n = 4, n=biological replicates.

**Techniques Used: **Mutagenesis, Immunofluorescence, Marker, Flow Cytometry, Staining

**Figure Legend Snippet:**Related to Fig. S2 A) A schematic for rescuing SFP in BRCA1 mutant cells by ablation of Smarcal1. B) DNA fiber analysis in a fork stalling assay with hydroxyurea (HU) on reprogramming day 5. At least 120 DNA fibers were measured per genotype. Analysis by one-way ANOVA. C) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cells lines. Data is shown as a ratio of dual allele targeting in each genotype relative to control. Statistical analysis with one-way ANOVA; wt ctrl n = 9, Brca1tr/+ n = 3, Brca1tr/tr n = 3, Brca1tr/tr Smarcal1−/− n = 3. D) Immunofluorescence and quantification for double strand break (DSB) marker phospho-H2AX (S139). Foci were counted on reprogramming day 5 (≥410 cells/genotype) and statistical analysis performed with one-way ANOVA. E) Immunofluorescence and quantification of ssDNA marker phospho-RPA(S33) on reprogramming day 5. Data collected from ≥140 cells per genotype and analyzed by one-way ANOVA. F) Cell proliferation analysis with CFSE on reprogramming day 5. Statistical analysis with one-way ANOVA.; wt ctrl n = 4, Smarcal1+/− n = 4, Smarcal1−/− n = 5, Brca1tr/tr n = 2, Brca1tr/tr Smarcal1−/− n = 3. G) Apoptosis analysis with Annexin V and propidium iodide (PI) on reprogramming day 5. Analysis by one-way ANOVA.; wt ctrl n = 3, Smarcal1+/− n = 3, Smarcal1−/− n = 3, Brca1tr/tr n = 3, Brca1tr/tr Smarcal1−/− n = 3. H) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. The number of AP positive colonies is shown as a ratio to wild type. Data analysis with one-way ANOVA.; wt ctrl n = 7, Brca1tr/+ n = 6, Smarcal1+/− n = 4, Smarcal1−/− n = 2, Brca1tr/tr n = 12, Brca1tr/tr Smarcal1+/− n = 4, Brca1tr/tr Smarcal1−/− n = 4. n= biological replicates.

**Techniques Used: **Mutagenesis, CRISPR, Control, Immunofluorescence, Marker, Staining

**Figure Legend Snippet:**Related to Fig. S3. A) A schematic for rescuing HDR in Brca1 mutant cells by ablation of 53bp1. Relevant substrates that can be repaired by HDR or NHEJ are shown. B) CRISPR/Cas9 based HDR assay with induced pluripotent stem (iPS) cells lines. Data is shown as a ratio of dual allele targeting in each genotype to dual allele targeting in the control. Statistical analysis using one-way ANOVA, except for the comparison between wt ctrl and 53bp1−/−, evaluated with a two-tailed, unpaired student’s t-test.; wt ctrl n = 9, Brca1tr/+ n = 3, 53bp1−/− n = 5, Brca1tr/tr n = 3, Brca1tr/tr53bp1+/− n = 2, Brca1tr/tr53bp1−/− n = 4. C) DNA fiber analysis in a fork stalling assay with hydroxyurea (HU) on reprogramming day 5. At least 120 DNA fibers were measured per genotype and data was analyzed by one-way ANOVA. D) Immunofluorescence and quantification for phospho-H2AX (S139). Foci counted on reprogramming day 5 (≥410 cells/genotype), statistical analysis performed with one-way ANOVA. E) Immunofluorescence and quantification of ssDNA marker phospho RPA(S33) on reprogramming day 5. ≥140 cells per genotype, analyzed by one-way ANOVA. For control and BRCA1, images are identical for Fig. 3 and Fig. 4 for panels c and d. F) Cell proliferation analysis with CFSE dye on reprogramming day 5. Statistics with one-way ANOVA.; wt ctrl n = 4,, 53bp1’/- n = 5, 53bp1−/− n = 3, Brca1tr/tr n = 2, Brca1tr/trSmarcal1−/− n = 3, Brca1tr/tr 53bp1−/− n = 3. G) Apoptosis analysis with Annexin V and propidium iodide (PI) on reprogramming day 5, analyzed by one-way ANOVA.; wt ctrl n = 3, 53bp1+/− n = 3, 53bp1−/− n = 2, Brca1tr/tr n = 3, Brca1tr/tr 53bp1−/− n = 3. H) Alkaline phosphatase (AP) staining and reprogramming efficiency quantification. Number of AP positive colonies is shown as a ratio to wild type, analyzed with one-way ANOVA.; wt ctrl n = 7, Brca1tr/+ n = 6, 53bp1+/− n = 7, 53bp1−/− n = 8, Brca1tr/tr n = 12, Brca1tr/tr 53bp1−/− n = 7. I) Quantification of Nanog positive colonies in the indicated genotypes, analyzed with one-way ANOVA.; n = 3 for each genotype. J) AP staining and reprogramming efficiency quantification in human 1023 fibroblasts from adult skin biopsy in control and 53BP1 knockdown (KD) conditions. Cells were fixed on day 25 post reprogramming factor transduction, statistical analysis using an unpaired, two-tailed student’s t-test.; ctrl n = 16, 53BP1 KD n = 16. Samples with n=2 are not used for statistical comparisons. All numbers indicated are biological replicates.

**Techniques Used: **Mutagenesis, CRISPR, Control, Comparison, Two Tailed Test, Immunofluorescence, Marker, Staining, Knockdown, Transduction

**Figure Legend Snippet:**KEY RESOURCES TABLE

**Techniques Used: **Purification, Transduction, Recombinant, Knock-Out, Plasmid Preparation, Software, Imaging

# mouse monoclonal anti phospho histone h2a x ser139 antibody (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139 Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139 antibody/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "TDP1 phosphorylation by CDK1 in mitosis promotes MUS81-dependent repair of trapped Top1-DNA covalent complexes"

**Article Title: **TDP1 phosphorylation by CDK1 in mitosis promotes MUS81-dependent repair of trapped Top1-DNA covalent complexes

**Journal: **The EMBO Journal

**doi: **10.1038/s44318-024-00169-3

**Figure Legend Snippet:**( A ) A schematic representation of the protocol for synchronization of MCF7 cells followed by chromatin immunoprecipitation (ChIP) with indicated antibodies at the CFS and non-CFS loci. ( B , C ) Genomic organization of the FRA3B and FRA16D regions, along with the primer sets of the distal (FDR) and central (FCR) region within the FRA3B locus; FRA16D locus, is designated. ( D – F ) Endogenous Top1 but not TDP1 preferentially localizes to CFSs upon CPT (15 nM; 24 h) treatment during mitosis. Quantification of cross-linked FRA3B-FCR, FRA3B-FDR and FRA16D loci chromatin-immunoprecipitated from MCF7 cells using the specified antibodies (Top1 and TDP1). The DSB marker γH2AX antibody was used as a positive control for FRA3B and FRA16D enrichment post CPT treatment. Fold enrichment over IgG was determined and is shown for each primer pair for the ChIP. Data are mean ± SD, n = 3 biological replicates. ns, non-significant ( P > 0.05); ** P ≤ 0.01 (one-way ANOVA). ( G – I ) Quantification of cross-linked FRA3B-FCR, FRA3B-FDR, and FRA16D loci chromatin-immunoprecipitated from MCF7 cells transfected with FLAG-TDP1 variants (WT or S61A) using the specified antibodies (endogenous Top1). Fold enrichment over IgG was determined and is shown for each primer pair for the ChIP. ( J – L ) Quantification of cross-linked FRA3B-FCR, FRA3B-FDR, and FRA16D loci chromatin-immunoprecipitated from MCF7 cells ectopically expressing the FLAG-TDP1 variants (WT and S61A) using the specified antibodies with or without CPT treatment (15 nM; 24 h). The DSB marker γH2AX antibody was used as a positive control for FRA3B and FRA16D enrichment post CPT treatment. Fold enrichment over IgG was determined and is shown for each primer pair for the ChIP. Data are mean ± SD, n = 3 biological replicates. ns, non-significant ( P > 0.05); ** P ≤ 0.01 (one-way ANOVA). ( M – P ) Quantification of cross-linked β-actin, GAPDH, β2-microglobulin and β-tubulin loci chromatin-immunoprecipitated from MCF7 cells ectopically expressing the FLAG-TDP1 variants (WT and S61A) using the specified antibodies with or without CPT treatment (15 nM; 24 h). The DSB marker γH2AX antibody was used as a positive control for β-actin, GAPDH, β2-microglobulin and β-tubulin enrichment post CPT treatment. Fold enrichment over IgG was determined and is shown for each primer pair for the ChIP. Data are mean ± SD, n = 3 biological replicates. n.s. non-significant ( P > 0.05); * P < 0.05, ** P < 0.01; (one-way ANOVA). .

**Techniques Used: **Chromatin Immunoprecipitation, Immunoprecipitation, Marker, Positive Control, Transfection, Expressing

**Figure Legend Snippet:**( A ) TDP1 −/− MEFs complemented with FLAG-TDP1 variants (TDP1 −/−/WT ; TDP1 −/−S61A ) or Empty vector (TDP1 −/−/EV ) were synchronized in mitosis and treated with or without CPT (10 μM, 1 h), released in presence of nocodazole followed by immunocytochemistry with anti-FLAG (red) to detect FLAG-TDP1 and anti-γH2AX (green) antibody. Cells were counterstained with DAPI to visualize mitotic nuclei (blue). ( B ) Quantification for the number of γH2AX foci per mitotic nucleus calculated for 50 nuclei. Data are mean ± SD, n = 3 biological replicates. * P ≤ 0.05; **** P ≤ 0.0001 (one-way ANOVA). ( C ) Representative merged image showing anaphase nucleus (blue) with γH2AX (green) foci on bridges resulting from CPT treatment in TDP1 −/−S61A MEFs. The enlarged view of the anaphase bridge has been shown. ( D ) Quantification of γH2AX-positive anaphase bridges in TDP1 −/−/EV ; TDP1 −/−/WT and TDP1 −/−S61A MEFs. Data are mean ± SD, n = 3 biological replicates. * P ≤ 0.05; ** P ≤ 0.01 (one-way ANOVA). ( E ) Representative merged image showing G1 primary nucleus (PN) with micronuclei (MN) harboring γH2AX (green) foci resulting from CPT treatment in mitosis. The enlarged view of the MN with γH2AX in TDP1 −/−S61A MEFs has been shown. ( F ) Quantification of the number of γH2AX-positive G1-MN in TDP1 −/−/EV ; TDP1 −/−/WT , and TDP1 −/−/S61A MEFs as indicated. Data are mean ± SD, n = 3 biological replicates. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 (one-way ANOVA). ( G ) TDP1 −/−/EV ; TDP1 −/−/WT , and TDP1 −/−/S61A MEFs were treated with CPT (15 nM; 9 h) in late S-phase as indicated to generate replication stress, synchronized in G2/M-phase followed by immunocytochemistry with anti-γH2AX (red) and anti-Top1cc (green) antibody. Cells were counterstained with DAPI to visualize mitotic nuclei (blue). ( H , I ) Quantifications for the number γH2AX foci ( H ) and Top1cc-positive γH2AX foci ( I ) per mitotic nucleus of TDP1 −/−/EV ; TDP1 −/−/WT and TDP1 −/−/S61A MEFs treated with CPT in late S calculated for 50 cells. Data are mean ± SD, n = 3 biological replicates. ns non-significant ( P > 0.05); * P ≤ 0.05; **** P ≤ 0.0001 (one-way ANOVA). Scale bars, 10 μm. .

**Techniques Used: **Plasmid Preparation, Immunocytochemistry

**Figure Legend Snippet:**( A – D ) TDP1 −/− MEFs complemented with EV or FLAG-TDP1 variants (TDP1 −/−/WT and TDP1 −/−/S61A ) were treated with or without CPT (15 nM; 24 h) or Aph (0.4 μM, 24 h) alone or in combination (CPT + APH 24 h) and enriched at M-phase. Representative images show break-induced repair with newly synthesized mitotic DNA marked by BrdU foci (red). The γH2AX foci, signifying the DNA strand breaks, are shown in green. Cells were counterstained with DAPI to visualize nuclei (blue). ( E , F ) Quantification of γH2AX foci per nucleus and percentage of BrdU-positive mitotic nucleus obtained from immunofluorescence by confocal microscopy were calculated for 20–25 cells. Data are mean ± SD, n = 3 biological replicates. ns, non-significant ( P > 0.05); *** P ≤ 0.001; **** P ≤ 0.0001 (one-way ANOVA). ( G ) Chromatin fractions were prepared from TDP1 −/−/WT and TDP1 −/−/S61A MEFs after CPT treatment (15 nM; 24 h) and analyzed by western blotting to detect FLAG-TDP1 variants and MUS81 with anti-FLAG and anti-MUS81 antibodies, respectively. Anti-HH3 and anti-pHH3 were used as chromosomal and mitotic markers. Protein levels of MUS81, FLAG-TDP1 WT , and FLAG-TDP1 S61A were analyzed in whole-cell lysates (WCE) to ensure equal levels of protein before chromatin fractionation. ( H ) Quantification showing the relative chromosomal enrichment of FLAG-TDP1 variants (WT or S61A) in interphases (Asn) and mitotic (M) chromosomes (left panel). Quantification showing the relative enrichment of MUS81 on mitotic chromosomes in interphases (Asn) and mitosis (M) for indicated cells treated with CPT (15 nM; 24 h) (right panel) n = 3 biological replicates. Data are mean ± SD, n = 3 biological replicates. ns, non-significant ( P > 0.05); * P ≤ 0.05; ** P ≤ 0.01 (one-way ANOVA). ( I ) Representative images of immunofluorescence microscopy showing induction of CPT (15 nM, 24 h)-induced γH2AX (green) and MUS81 (red) foci during mitosis in TDP1 −/−/EV , TDP1 −/−/WT , and TDP1 −/−/S61A MEFs, co-transfected either with Si Ctrl or Si MUS81 to knockdown MUS81. Cells were counterstained with DAPI to visualize mitotic nuclei (blue). Note: Colocalization of CPT-induced γH2AX and MUS81 foci in merged images indicates MUS81 overloading amplifies mitotic DNA breaks in TDP1 −/−/S61A MEFs. ( J ) Representative western blot analysis for the immunofluorescence microscopy in ( I ) to detect the levels of MUS81 and FLAG-TDP1 in TDP1 −/−/EV , TDP1 −/−/WT , and TDP1 −/−/S61A MEFs, co-transfected either with Si Ctrl or Si MUS81 to knockdown MUS81. β-actin has been used as loading control. ( K ) Quantifications of MUS81 foci on the mitotic chromosomes scored for 50 nuclei (each category) as depicted by the corresponding bar diagram. Data are mean ± SD, n = 3 biological replicates. ns, non-significant ( P > 0.05); ** P ≤ 0.01 (one-way ANOVA). ( L ) Quantifications for CPT-induced γH2AX foci on mitotic nuclei in indicated cells ( n = 50 cells from three biological replicates; mean ± SD). Note: siRNA knockdown of MUS81 significantly reduced CPT-induced γH2AX. ns, non-significant ( P > 0.05); ****P ≤ 0.0001 (one-way ANOVA). ( M ) A schematic representation for the protocol followed for the ChIP of endogenous MUS81 at the CFS loci in mitosis following CPT (15 nM, 24 h) treatment. ( N – P ) Quantification of cross-linked FRA3B-FCR, FRA3B-FDR, and FRA16D loci chromatin-immunoprecipitated from MCF7 cells transfected with empty vector (EV) or FLAG-TDP1 variants (WT or S61A) using the specified antibodies. Fold enrichment over IgG was determined and is shown for each primer pair for the ChIP. Data are mean ± SD, n = 3 biological replicates. * P ≤ 0.1; ** P ≤ 0.01 (one-way ANOVA). Scale bars, 10 μm. .

**Techniques Used: **Synthesized, Immunofluorescence, Confocal Microscopy, Western Blot, Fractionation, Microscopy, Transfection, Knockdown, Control, Immunoprecipitation, Plasmid Preparation

**Figure Legend Snippet:**Reagents and tools table

**Techniques Used: **Virus, Recombinant, Transfection, Modification, Protease Inhibitor, Reverse Transcription, Mutagenesis, Imaging, Software

# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "Crucial role of the NSE1 RING domain in Smc5/6 stability and FANCM-independent fork progression"

**Article Title: **Crucial role of the NSE1 RING domain in Smc5/6 stability and FANCM-independent fork progression

**Journal: **Cellular and Molecular Life Sciences: CMLS

**doi: **10.1007/s00018-024-05275-3

**Figure Legend Snippet:**nse1 mutants show increased levels of DNA-damage and genomic instability. A. Up, representative images of aberrant anaphases; bottom, frequency of aberrant anaphases (anaphase bridges and lagging chromosomes), relative to the total number of anaphases scored; a minimum of 80 anaphases were counted per experiment; bars indicate means and error bars SD. B. Frequency of micronuclei; bars indicate means and error bars SD. C . Western blot analysis of the indicated Smc5/6 subunits and different markers of endogenous DNA damage (γH2AX) and checkpoint activation (phospho (S345) CHK1, phospho (T48) CHK2) in HEK293T cells. D . Western blot analysis of DNA damage checkpoint activation markers in wild type and nse1-KO MRC5-VI cells. Where indicated, cells were treated with etoposide 5 µM for 16 h to induce DNA damage. E . Left: Clonogenic assay of HEK293T nse1-ΔR cells expressing (+) or not (-) NSE1 after acute exposure (2 h) to 250 µM MMS. Right: quantification of colony intensity from three independent experiments. Dots are individual clonogenic assay measures. Error bars are SD values. In A, B and E: ns, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by a one-way ANOVA followed by Tukey’s multiple comparison test

**Techniques Used: **Western Blot, Activation Assay, Clonogenic Assay, Expressing, Comparison

# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "Integrative modeling uncovers p21-driven drug resistance and prioritizes therapies for PIK3CA-mutant breast cancer"

**Article Title: **Integrative modeling uncovers p21-driven drug resistance and prioritizes therapies for PIK3CA-mutant breast cancer

**Journal: **NPJ Precision Oncology

**doi: **10.1038/s41698-024-00496-y

**Figure Legend Snippet:**a β-gal senescence assay of parental and RP cells left untreated or treated with 1 μM BYL719 for 48 h. Percentages of β-gal positive cells were calculated by dividing the number of β-gal (blue) cells over the total number of nuclei (DAPI). Violin plots showing median plus 1st and 3 rd quartiles. Kruskal–Wallis analysis, * p < 0.05, ** p < 0.01, **** p < 0.0001; #### p < 0.0001 for comparison between cells in BYL719. Right, β-gal stains (bright-field) of parental and RP cells treated as indicated. Scale bar = 50 μm ( n = 10 fields of view). b Percentage of senescence in RPs with targeted p21 KD, quantified as in ( a ). Right, β-gal staining of T47D cells treated as indicated. Violin plot showing median plus 1st and 3rd quantiles. Kruskal–Wallis analysis, * p < 0.05, **** p < 0.0001; #### p < 0.0001 for comparison between RPs and Parental cells ( n > 10 fields of view). c γH2AX fluorescence intensity was quantified in parental T47D cells and RPs upon 1 μM BYL719 treatment for 48 h; each data point represents one nucleus. Mean ± SD is shown, one-way ANOVA, **** p < 0.0001 ( n > 40 nuclei per group). Bottom, representative γH2AX images of parental and RPs in 1 μM BYL719 for 48 h. Scale bar = 5 μm. d 53BP1 fluorescence intensity was quantified in parental T47D cells and RPs upon 1 μM BYL719 treatment for 48 h; each data point represents one nucleus. Mean ± SD is shown, one-way ANOVA, **** p < 0.0001. Bottom, representative 53BP1 images of parental and RPs in 1 μM BYL719 for 48 h. Scale bar = 5 μm. e DNA damage quantification of T47D cells with targeted p21 KD and treated with 1 μM BYL719 for 48 h. γH2AX fluorescence intensity was quantified as in ( d ). Mean ± SD is shown, one-way ANOVA, **** p < 0.0001 ( n > 40 nuclei per group). Representative images of γH2AX in parental and RPs. Scale bar =5 μm. f WB analysis of DNA damage repair activators in T47D parental and RP cells treated with 1 μM BYL719 for 48 h.

**Techniques Used: **Comparison, Staining, Fluorescence

**Figure Legend Snippet:**a WB of parental and RPs treated with 1 μM BYL719 for 24 h and probed with the indicated antibodies. b T47D parental cells and RPs were treated with 1 μM BYL719 alone or in combination with either 5 or 10 μM of the CHK1 inhibitor MK-8776 for 3 days. Cell viability was quantified using CellTiter-glo, Data presented as mean ± SD, one-way ANOVA, **** p < 0.0001 ( n = 3). c T47D parental cells and BYL719-RPs were treated with the indicated doses of MK-8776 for 4 days and percentage of dead cells quantified through PI staining. Data presented as mean ± SD, one-way ANOVA comparison between DMSO and MK-8776 treatments of the same cell line, ** p < 0.01. **** p < 0.0001 ( n = 3). d Growth curve of T47D cells treated with BYL719 (1 μM) or MK-8776 (5 μM) alone or in combination for 6 days. Drugs were refreshed every 2 days. Data points represent mean ± SD, one-way ANOVA, **** p < 0.0001, Student’s t test, ### p < 0.001 comparing response to BYL719 alone or in combination with MK-8776 between RPs and parental cells ( n = 3). e DNA damage quantification in parental and RPs treated with 1 μM BYL719 and 5 μM MK-8776 for 48 h. γH2AX fluorescence intensity was quantified and each data point represents one nucleus. Mean ± SD is shown, one-way ANOVA, *** p < 0.001 ( n > 40 nuclei per group). f β-gal assay of parental and RPs treated with BYL719 (1 μM) or MK-8776 (5 μM) alone or in combination for 4 days. Percentage of positive cells was calculated as the number of β-gal positive cells over total number of nuclei. Violin plot showing median plus 1st and 3rd quantiles. Kruskal–Wallis analysis, * p < 0.05; # p < 0.05, ## p < 0.01, #### p < 0.0001 for comparison between treatments of the same cells ( n > 10 fields of view from each triplicate wells). g Cell death assay of T47D parental cells and RPs treated with BYL719 (1 μM) or MK-8776 (5 μM) alone or in combination for 6 days. Number of dead cells were quantified through PI staining and normalized over total cell number quantified by Hoechst 33342 dye. Data are presented as mean ± SEM, one-way ANOVA, *** p < 0.001, **** p < 0.0001; # p < 0.05, #### p < 0.0001 for comparison with BYL719 treated cells ( n = 3 replicates of culture). Inserts show PI-stained (red) T47D parental cells and RP1 cells in different conditions. Nuclei counterstained with Hoechst33342 (blue). Scale bar = 100 μm. h BYL719 treatments cause DNA damage and senescence in T47D cells. Selection of high-p21 levels promotes repair of damaged DNA and evasion of drug-induced cellular senescence. p21 can also promote formation of the cyclin D1-CDK4/CDK6 complex, further supporting cell cycle progression , . In combination with BYL719, the CHK1 inhibitor MK-8776 leads to excessive DNA damage and death of BYL719-resistant cells.

**Techniques Used: **Staining, Comparison, Fluorescence, Selection

**Figure Legend Snippet:**

**Techniques Used: **

# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99

## Images

### 1) Product Images from "Roles of USP1 in Ewing sarcoma"

**Article Title: **Roles of USP1 in Ewing sarcoma

**Journal: **Genes & Cancer

**doi: **10.18632/genesandcancer.235

**Figure Legend Snippet:**( A ) A673 cells were sorted into the CD133 high and CD133 low populations. Each cell population was analyzed for the sensitivity to indicated concentration of cisplatin or doxorubicin using the IncuCyte live-cell imaging system. ( B ) The protein levels of the mediators of DNA damage response in the CD133 high and CD133 low populations. The CD133 high and CD133 low populations derived from A673 cells were left untreated or treated with 0.01 μM cisplatin for the indicated time and the levels of phosphorylated ATM, ATR, CHK1, CHK2, and H2AX as well as the levels of SLFN11 were examined by immunoblotting. Tubulin serves as a loading control. ( C ) The CD133 low population displays increased FANCD2 mono-ubiquitination and reduced USP1 levels compared with the CD133 high population. The CD133 high and CD133 low populations derived from A673 cells were left untreated or treated with 0.01 μM cisplatin for the indicated time and the levels of FANCD2 mono-ubiquitination and USP1 were examined by immunoblotting. Tubulin serves as a loading control.

**Techniques Used: **Concentration Assay, Live Cell Imaging, Derivative Assay, Western Blot

# mouse monoclonal anti phospho histone h2a x ser139 (Millipore)

## Structured Review

Mouse Monoclonal Anti Phospho Histone H2a X Ser139, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more

https://www.bioz.com/result/mouse monoclonal anti phospho histone h2a x ser139/product/Millipore

Average 86 stars, based on 1 article reviews

Price from $9.99 to $1999.99