Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)

Rabbit Monoclonal H2A.X phospho S139 antibody. Carrier free. Suitable for ICC/IF, IP, Dot, WB, IHC-P and reacts with Human, Synthetic peptide samples. Cited in 15 publications.

Be the first to review this product! Submit a review

Related conjugates and formulations

ab81299
Unconjugated
Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]
ab206898
Conjugated
Alexa Fluor® 594 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]
ab195188
Conjugated
Alexa Fluor® 488 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]
ab195189
Conjugated
Alexa Fluor® 647 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]
ab195190
Conjugated
HRP Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]
ab206900
Conjugated
Alexa Fluor® 555 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y]

Images

Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (AB215967), expandable thumbnail

Publications

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	ICC/IF	IP	Dot	WB	IHC-P
Human	Tested	Tested	Expected	Tested	Tested
Mouse	Predicted	Predicted	Predicted	Predicted	Not recommended
Rat	Predicted	Predicted	Predicted	Predicted	Not recommended
Sheep	Predicted	Predicted	Predicted	Predicted	Not recommended
Synthetic peptide	Not recommended	Not recommended	Tested	Not recommended	Not recommended

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Mouse, Rat, Sheep	Dilution info -	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Synthetic peptide	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Mouse, Rat, Sheep	Dilution info -	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Synthetic peptide	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Synthetic peptide	Dilution info -	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Human	Dilution info Use at an assay dependent concentration.	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Mouse, Rat, Sheep	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Mouse, Rat, Sheep	Dilution info -	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Synthetic peptide	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Mouse, Rat, Sheep, Synthetic peptide	Dilution info -	Notes -

Associated Products

Select an associated product type

6 products for Alternative Product

5 products for Alternative Version

Target data

See full target information H2AX phospho S139

Function

Variant histone H2A which replaces conventional H2A in a subset of nucleosomes. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. Required for checkpoint-mediated arrest of cell cycle progression in response to low doses of ionizing radiation and for efficient repair of DNA double strand breaks (DSBs) specifically when modified by C-terminal phosphorylation.

Alternative names

H2AFX, H2AX, Histone H2AX, H2a/x, Histone H2A.X, H2AS139p, H2AXS139p, H2A.XS139p

Recommended products

Rabbit Monoclonal H2A.X phospho S139 antibody. Carrier free. Suitable for ICC/IF, IP, Dot, WB, IHC-P and reacts with Human, Synthetic peptide samples. Cited in 15 publications.

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Carrier free: Yes
Clone number: EP854(2)Y
Purification technique: Affinity purification Protein A
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: +4°C
Storage information: Do Not Freeze

Notes

ab215967 is the carrier-free version of Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299.

Patented technology
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

What are the advantages of a recombinant monoclonal antibody?
This product is a recombinant monoclonal antibody, which offers several advantages including:

- High batch-to-batch consistency and reproducibility
- Improved sensitivity and specificity
- Long-term security of supply
- Animal-free batch production

For more information, read more on recombinant antibodies.

Conjugation ready
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

Compatibility
This product is compatible with the Maxpar^® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar^® is a trademark of Fluidigm Canada Inc.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.

Activity summary

Gamma H2A.X also known as phospho H2A.X or γH2A.X is a phosphorylated form of the histone variant H2A.X. It has a molecular weight of about 14 kilodaltons and occurs primarily in they nucleus. When DNA double-strand breaks (DSBs) occur serine 139 in H2A.X undergoes rapid phosphorylation resulting in gamma H2A.X. This modification happens swiftly at the site of damage and gamma H2A.X spreads over a large chromatin area facilitating the recruitment of DNA repair proteins. Gamma H2A.X staining typically evaluated using gamma H2A.X immunofluorescence techniques aids in identifying the presence and extent of DNA damage.

Biological function summary

Gamma H2A.X plays a role in DNA damage response and repair. It does not operate alone; it acts as part of a complex with other repair proteins. The formation of gamma H2A.X foci at DNA damage sites creates a signal attracting repair factors that help maintain genome stability. Its interaction with MDC1 and ATM proteins exemplifies its significant role in orchestrating an effective response to DNA damage. Beyond DNA repair gamma H2A.X influences cell cycle checkpoints permitting cells to pause and repair before proceeding with division.

Pathways

Gamma H2A.X plays a pivotal role in the DNA damage response (DDR) pathway. This pathway is essential for detecting and repairing DNA lesions to uphold genomic integrity. Within the DDR pathway gamma H2A.X is closely associated with proteins such as NBS1 and BRCA1 which assist in repairing double-strand breaks. In addition gamma H2A.X is integral to the ATM-ATR signaling pathway where its activation promotes cell survival following genotoxic stress by signaling for damage repair or triggering apoptosis.

Associated diseases and disorders

Gamma H2A.X has connections to cancer and neurodegeneration. Aberrant DNA repair pathways often indicated by persistent gamma H2A.X signals correlate with tumor formation and progression. For instance a failure to repair DNA damage effectively can lead to mutations that drive cancer development. Gamma H2A.X also links to neurodegenerative diseases where dysregulated DNA repair contributes to neuronal cell death. Proteins like p53 which regulate cell cycle and apoptosis further connect to gamma H2A.X bridging its role in disease pathogenesis.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

14 product images

Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Immunofluorescence staining of A549 (Human lung carcinoma cell line) labeling gamma H2A.X (phospho S139) (green) with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299.
Cells were fixed in 4% paraformaldehyde for 15 minutes and permeabilized in PBS-0.2% Triton for 10 minutes. After blocked for 1 hour, primary antibody was diluted in blocking buffer (1/100) and incubated with fixed cells overnight at 4°C. Cells were washed and incubated with secondary antibodies (1/100) for 1 hour at room temperature. All slides were counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Immunofluorescence was performed using confocal laser scanning microscopy (Lecia) or fluorescence microscopy (Olympus).
Dexamethasone, DEX; Cisplatin, DDP.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Clone EP854(2)Y (ab215967) has been successfully conjugated by Abcam. This image was generated using Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (Alexa Fluor® 488). Please refer to Alexa Fluor® 488 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195188 for protocol details.
Alexa Fluor® 488 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195188 staining Histone H2A.X in Jurkat cells. The cells were incubated with 25uM ETP for 5 hours (Treated) or solvent-only for control purposes (Non-treated). The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 10% normal goat serum in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with Alexa Fluor® 488 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195188 at 1/50 dilution (shown in green) and Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 594), at 1/200 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at 1/40 immunoprecipitating Histone H2A.X (phospho S139) in HepG2 (human hepatocellular carcinoma epithelial) whole cell lysate observed at 15 KDa (lanes 1 and 2).
Lane 1 (input): HepG2 treated with etoposide and TSA whole cell lysate 10μg
Lane 2 (+): Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 + HepG2 treated with etoposide and TSA whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 in HepG2 treated with etoposide and TSA
For western blotting, Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 (Purified) at 1/200 dilution and VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366), was used for detection at 1/1000 dilution.
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
All lanes: Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299)
Predicted band size: 15 kDa
Exposure time: 30s
Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Immunocytochemistry/Immunofluorescence analysis of HeLa cells (untreated and treated with H₂O₂) labelling Histone H2A.X (phospho S139) with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at 1/250. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. The cells were co-stained with Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291, a mouse anti-tubulin (1/1000) using Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/1000) as the secondary antibody. Nuclei counterstained with DAPI (blue).
Control 1: primary antibody (1/250) and secondary antibody, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/1000).
Control 2: Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (1/1000) and secondary antibody, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Clone EP854(2)Y (ab215967) has been successfully conjugated by Abcam. This image was generated using Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (Alexa Fluor® 647). Please refer to Alexa Fluor® 647 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195189 for protocol details.
Alexa Fluor® 647 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195189 staining Histone H2A.X in Jurkat cells. The cells were incubated with 25uM ETP for 5 hours (Treated) or solvent-only for control purposes (Non-treated). The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 10% normal goat serum in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with Alexa Fluor® 647 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195189 at 1/200 dilution (shown in red) and Alexa Fluor® 488 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 488), at 1/200 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
This image is courtesy of a customer review submitted by Carl Hobbs.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 staining H2A.X in Human Testis tissue sections by Immunohistochemistry (Formalin/PFA fixed paraffin embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 10 minutes at 21°C; antigen retrieval was by heat mediation in Citric acid. Samples were incubated with primary antibody (1/50 in TBS) for 2 hours at 21°C. A biotin conjugated Anti-Rabbit IgG (goat polyclonal) was used as the secondary antibody at a 1/250 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Dot Blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Dot blot analysis of Histone H2A.X single phospho peptide pS139 (lane 1) and Histone H2A.X non-phospho peptide (lane 2) with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at 1/1000. Blocking and diluting buffer was 5% NFDM/TBST. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) ab97051 Peroxidase conjugated Goat Anti-Rabbit IgG, (H+L) at 1/100,000.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Immunohistochemical staining of paraffin embedded human brain with unpurified Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Immunohistochemical staining of paraffin embedded human brain with purified Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at a working dilution of 1 in 100. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
Immunohistochemical analysis of formalin/PFA-fixed paraffin-embedded human kidney transitional cell carcinoma using unpurified Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at a dilution of 1/100.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
This data was developed using the same antibody clone in a different buffer formulation (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299). Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue labeling gamma H2A.X with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at 1/3000 (0.352 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human breast carcinoma without lambda protein phosphatase treatment (image A). No signal was detected when tissues were treated with lambda protein phosphatase (image B). The section was incubated with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 for 15 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB ab209101. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.
Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299).
Mouse anti GAPDH was used as a GAPDH loading control.
All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299) at 1/1000 dilution
Lane 1: HepG2 Vehicle Control Etoposide (0 uM, 90 min) at 20 µg
Lane 2: HepG2 Etoposide treated (100 uM, 90 min) at 20 µg
Lane 3: Jurkat Vehicle Control Etoposide (0 uM, 90 min) at 20 µg
Lane 4: Jurkat Treated Etoposide (100 uM, 90 min) at 20 µg
Lane 5: Human Brain at 20 µg
Predicted band size: 15 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
This data was developed using the same antibody clone in a different buffer formulation (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299). Immunohistochemical analysis of paraffin-embedded Human cervical carcinoma tissue labeling gamma H2A.X with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 at 1/3000 (0.352 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human cervical carcinoma without lambda protein phosphatase treatment (image A). No signal was detected when tissues were treated with lambda protein phosphatase (image B). The section was incubated with Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 for 15 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB ab209101. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.
Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free (ab215967)
This data was developed using Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299, the same antibody clone in a different buffer formulation.
Western blot: Rabbit Monoclonal [EP854(2)Y] to gamma H2A.X (phospho S139) Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 15 kDa in HepG2 Vehicle Control Etoposide (0 uM, 90 min) cell lysates with no signal observed at this size in HepG2 Etoposide treated (100 uM, 90 min) cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc BSA in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.
All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab81299) at 1/1000 dilution
Lane 1: HepG2 Vehicle Control Etoposide (0 uM, 90 min) at 20 µg
Lane 2: HepG2 Etoposide treated (100 uM, 90 min) at 20 µg
Lane 3: Jurkat Vehicle Control Etoposide (0 uM, 90 min) at 20 µg
Lane 4: Jurkat Treated Etoposide (100 uM, 90 min) at 20 µg
Lane 5: Human Brain at 20 µg
Secondary
All lanes: Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 15 kDa
Observed band size: 15 kDa