Anti-EZHIP antibody [EPR28396-24] (ab316856)

Rabbit Recombinant Monoclonal EZHIP antibody. Suitable for IHC-P, ICC/IF, WB, Flow Cyt (Intra), IP and reacts with Human samples.

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Images

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EZHIP antibody [EPR28396-24] (AB316856), expandable thumbnail

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal

Immunogen

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

Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	IHC-P	ICC/IF	WB	Flow Cyt (Intra)	IP
Human	Tested	Tested	Tested	Tested	Tested
Mouse	Not recommended	Not recommended	Not recommended	Not recommended	Not recommended
Rat	Not recommended	Not recommended	Not recommended	Not recommended	Not recommended

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/500	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Not recommended
Not recommended

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

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/100	Notes -

Not recommended
Not recommended

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

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/1000	Notes -

Not recommended
Not recommended

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

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/500	Notes -

Not recommended
Not recommended

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

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/30	Notes -

Not recommended
Not recommended

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

Target data

See full target information EZHIP

Function

Inhibits PRC2/EED-EZH1 and PRC2/EED-EZH2 complex function by inhibiting EZH1/EZH2 methyltransferase activity, thereby causing down-regulation of histone H3 trimethylation on 'Lys-27' (H3K27me3) (PubMed:29909548, PubMed:30923826, PubMed:31086175, PubMed:31451685). Probably inhibits methyltransferase activity by limiting the stimulatory effect of cofactors such as AEBP2 and JARID2 (PubMed:30923826). Inhibits H3K27me3 deposition during spermatogenesis and oogenesis (By similarity).

Alternative names

CXorf67, EZHIP, EZH inhibitory protein

Recommended products

Rabbit Recombinant Monoclonal EZHIP antibody. Suitable for IHC-P, ICC/IF, WB, Flow Cyt (Intra), IP and reacts with Human samples.

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Clone number: EPR28396-24
Purification technique: Affinity purification Protein A
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage duration: 1-2 weeks
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: -20°C
Aliquoting information: Upon delivery aliquot
Storage information: Avoid freeze / thaw cycle

Notes

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

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.

Supplementary info

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

Activity summary

The EZHIP (EZH Inhibitory Protein) also known as CATACOMB is a protein that inhibits the activity of EZH2 an important enzyme in the PRC2 (Polycomb Repressive Complex 2). EZHIP is a protein with a molecular mass of approximately 90 kDa. It expresses in various tissues with notable presence in the male and female reproductive tracts as well as in several cancer cell lines. The protein's role involves modulating chromatin structure by interfering with methyltransferase activity affecting gene silencing.

Biological function summary

EZHIP interacts with the Polycomb Repressive Complex 2 (PRC2) altering its methylation activity on histone H3 at lysine 27 (H3K27me3) an important mark for epigenetic gene silencing. By engaging the complex EZHIP hinders the catalytic efficiency of EZH2 reducing the repression of target genes. This interaction significantly impacts epigenetic regulation as alterations to methylation patterns influence cellular functions and differentiation processes.

Pathways

EZHIP's function affects pathways related to chromatin modification and epigenetic regulation. It is central to the regulation of the JAK-STAT signaling pathway and interacts closely with EZH2 and possibly other members of the Polycomb group proteins. EZHIP shapes the chromatin landscape influencing the transcriptional output by collaborating or competing with other transcription factors and modifiers within these pathways.

Associated diseases and disorders

EZHIP's aberrant expression is linked to cancers including gliomas and germ cell tumors. The relationship between EZHIP and EZH2 becomes important as alterations in EZH2 activity lead to abnormal gene silencing associated with oncogenesis. Additionally research indicates that dysregulation involving EZHIP might contribute to developmental disorders through improper epigenetic marks. Further study about its interactions could provide insights into therapeutic approaches for conditions linked to epigenetic misregulation.

Product promise

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Full details and terms and conditions can be found here:
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9 product images

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling EZHIP with ab316856 at 1/500 (0.996 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Negative control: No staining on human liver.

The section was incubated with ab316856 for 30 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 LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins
Western blot - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Negative control: A204.
The molecular weight observed is consistent with what has been described in the literature (PMID: 34880314).
Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.
In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining at 1/200000 dilution.
All lanes: Western blot - Anti-EZHIP antibody [EPR28396-24] (ab316856) at 1/1000 dilution
Lane 1: U-2OS (human bone osteosarcoma epithelial cell) whole cell lysate at 50 µg with 5% NFDM/TBST
Lane 2: A-204 (human muscle rhabdomyosarcoma cell) whole cell lysate at 50 µg with 5% NFDM/TBST
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Observed band size: 40 kDa, 75 kDa, 36 kDa
Exposure time: 10s
Western blot - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Low expression: 293T (PMID: 34880314).
The molecular weight observed is consistent with what has been described in the literature (PMID: 34880314).
In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining at 1/200000 dilution.
All lanes: Western blot - Anti-EZHIP antibody [EPR28396-24] (ab316856) at 1/1000 dilution
Lane 1: U-2OS (human bone osteosarcoma epithelial cell) whole cell lysate at 50 µg with 5% NFDM/TBST
Lane 2: 293T (human embryonic kidney epithelial cell) whole cell lysate at 50 µg with 5% NFDM/TBST
Secondary
All lanes: Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution
Observed band size: 40 kDa, 75 kDa, 36 kDa
Exposure time: 6s
Immunoprecipitation - Anti-EZHIP antibody [EPR28396-24] (ab316856)
EZHIP was immunoprecipitated from 0.35 mg U-2 OS (human bone osteosarcoma epithelial cell) whole cell lysate with ab316856 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab316856 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
Lane 1: U-2 OS (human bone osteosarcoma epithelial cell) whole cell lysate

Lane 2: ab316856 IP in U-2 OS (human bone osteosarcoma epithelial cell) whole cell lysate

Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab316856 in U-2 OS whole cell lysate
Lysate was freshly made and used for Western blotting immediately to minimize protein degradation.
All lanes: Immunoprecipitation - Anti-EZHIP antibody [EPR28396-24] (ab316856) at 1/30 dilution
All lanes: U-2 OS (human bone osteosarcoma epithelial cell) whole cell lysate with 5% NFDM/TBST
Secondary
All lanes: Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/5000 dilution
Exposure time: 10s
Flow Cytometry (Intracellular) - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized 293T (human embryonic kidney epithelial cell, Left) / U-2 OS (human bone osteosarcoma epithelial cell, Right) cells labelling EZHIP with ab316856 at 1/500 dilution (0.1 ug)/Red compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).
Goat Anti-Rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/5000 dilution was used as the secondary antibody.
Low expression: 293T.
Immunocytochemistry/ Immunofluorescence - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized U-2 OS (human bone osteosarcoma epithelial cell) cells labelling EZHIP with ab316856 at 1/100 (4.98 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).
Confocal image showing nuclear staining in U-2 OS cell line.

Low expression: 293T (PMID: 34880314).

Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).
Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5ug/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling EZHIP with ab316856 at 1/500 (0.996 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Negative control: No staining on human kidney (PMID: 31451685).

The section was incubated with ab316856 for 30 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 LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Immunohistochemical analysis of paraffin-embedded Human placenta tissue labeling EZHIP with ab316856 at 1/500 (0.996 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Positive staining on human placenta (PMID: 27699219).

The section was incubated with ab316856 for 30 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 LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EZHIP antibody [EPR28396-24] (ab316856)
Immunohistochemical analysis of paraffin-embedded Human testis tissue labeling EZHIP with ab316856 at 1/500 (0.996 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Positive staining on human testis (PMID: 31451685).

The section was incubated with ab316856 for 30 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 LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins