Name: Anti-METTL3 antibody [EPR18810]
SKU: ab195352
Rating: 5 (8 reviews)

Anti-METTL3 antibody [EPR18810] (ab195352) is a rabbit monoclonal antibody detecting METTL3 in Western Blot, Flow Cytometry (Intra), Flow Cytometry, IP, IHC-P, ICC/IF. Suitable for Human, Mouse, Rat.

- Clone EPR18810 is the most cited clone to METTL3

- KO validated for confirmed specificity

- Biophysical QC for unrivalled batch-batch consistency

- Over 290 publications

Images

Immunohistochemistry - Anti-METTL3 antibody [EPR18810] (AB195352), expandable thumbnail

Publications

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

	IP	WB	ICC/IF	Flow Cyt (Intra)	IHC-P
Human	Tested	Tested	Tested	Tested	Tested
Mouse	Expected	Tested	Expected	Expected	Tested
Rat	Expected	Tested	Expected	Expected	Tested
Cat	Predicted	Predicted	Predicted	Predicted	Predicted
Cow	Predicted	Predicted	Predicted	Predicted	Predicted
Dog	Predicted	Predicted	Predicted	Predicted	Predicted
Goat	Predicted	Predicted	Predicted	Predicted	Predicted
Guinea pig	Predicted	Predicted	Predicted	Predicted	Predicted
Horse	Predicted	Predicted	Predicted	Predicted	Predicted
Pig	Predicted	Predicted	Predicted	Predicted	Predicted
Primates	Predicted	Predicted	Predicted	Predicted	Predicted
Sheep	Predicted	Predicted	Predicted	Predicted	Predicted

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 1/50	Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Sheep, Goat, Horse, Guinea pig, Cow, Cat, Dog, Pig, Primates	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/1000	Notes Milk recommended as blocking agent. Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Rat	Dilution info 1/1000	Notes Milk recommended as blocking agent. Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Species Human	Dilution info 1/1000	Notes Milk recommended as blocking agent.

Predicted
Predicted

Species	Dilution info	Notes
Species Sheep, Goat, Horse, Guinea pig, Cow, Cat, Dog, Pig, Primates	Dilution info -	Notes -

Tested
Tested

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

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Sheep, Goat, Horse, Guinea pig, Cow, Cat, Dog, Pig, Primates	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

Predicted
Predicted

Species	Dilution info	Notes
Species Sheep, Goat, Horse, Guinea pig, Cow, Cat, Dog, Pig, Primates	Dilution info -	Notes -

Tested
Tested

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

Predicted
Predicted

Species	Dilution info	Notes
Species Sheep, Goat, Horse, Guinea pig, Cow, Cat, Dog, Pig, Primates	Dilution info -	Notes -

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Target data

See full target information METTL3

Function

The METTL3-METTL14 heterodimer forms a N6-methyltransferase complex that methylates adenosine residues at the N(6) position of some RNAs and regulates various processes such as the circadian clock, differentiation of embryonic and hematopoietic stem cells, cortical neurogenesis, response to DNA damage, differentiation of T-cells and primary miRNA processing (PubMed:22575960, PubMed:24284625, PubMed:25719671, PubMed:25799998, PubMed:26321680, PubMed:26593424, PubMed:27281194, PubMed:27373337, PubMed:27627798, PubMed:28297716, PubMed:29348140, PubMed:29506078, PubMed:30428350, PubMed:9409616). In the heterodimer formed with METTL14, METTL3 constitutes the catalytic core (PubMed:27281194, PubMed:27373337, PubMed:27627798). N6-methyladenosine (m6A), which takes place at the 5'-[AG]GAC-3' consensus sites of some mRNAs, plays a role in mRNA stability, processing, translation efficiency and editing (PubMed:22575960, PubMed:24284625, PubMed:25719671, PubMed:25799998, PubMed:26321680, PubMed:26593424, PubMed:28297716, PubMed:9409616). M6A acts as a key regulator of mRNA stability: methylation is completed upon the release of mRNA into the nucleoplasm and promotes mRNA destabilization and degradation (PubMed:28637692). In embryonic stem cells (ESCs), m6A methylation of mRNAs encoding key naive pluripotency-promoting transcripts results in transcript destabilization, promoting differentiation of ESCs (By similarity). M6A regulates the length of the circadian clock: acts as an early pace-setter in the circadian loop by putting mRNA production on a fast-track for facilitating nuclear processing, thereby providing an early point of control in setting the dynamics of the feedback loop (By similarity). M6A also regulates circadian regulation of hepatic lipid metabolism (PubMed:30428350). M6A regulates spermatogonial differentiation and meiosis and is essential for male fertility and spermatogenesis (By similarity). Also required for oogenesis (By similarity). Involved in the response to DNA damage: in response to ultraviolet irradiation, METTL3 rapidly catalyzes the formation of m6A on poly(A) transcripts at DNA damage sites, leading to the recruitment of POLK to DNA damage sites (PubMed:28297716). M6A is also required for T-cell homeostasis and differentiation: m6A methylation of transcripts of SOCS family members (SOCS1, SOCS3 and CISH) in naive T-cells promotes mRNA destabilization and degradation, promoting T-cell differentiation (By similarity). Inhibits the type I interferon response by mediating m6A methylation of IFNB (PubMed:30559377). M6A also takes place in other RNA molecules, such as primary miRNA (pri-miRNAs) (PubMed:25799998). Mediates m6A methylation of Xist RNA, thereby participating in random X inactivation: m6A methylation of Xist leads to target YTHDC1 reader on Xist and promote transcription repression activity of Xist (PubMed:27602518). M6A also regulates cortical neurogenesis: m6A methylation of transcripts related to transcription factors, neural stem cells, the cell cycle and neuronal differentiation during brain development promotes their destabilization and decay, promoting differentiation of radial glial cells (By similarity). METTL3 mediates methylation of pri-miRNAs, marking them for recognition and processing by DGCR8 (PubMed:25799998). Acts as a positive regulator of mRNA translation independently of the methyltransferase activity: promotes translation by interacting with the translation initiation machinery in the cytoplasm (PubMed:27117702). Its overexpression in a number of cancer cells suggests that it may participate in cancer cell proliferation by promoting mRNA translation (PubMed:27117702). During human coronavirus SARS-CoV-2 infection, adds m6A modifications in SARS-CoV-2 RNA leading to decreased RIGI binding and subsequently dampening the sensing and activation of innate immune responses (PubMed:33961823).

Alternative names

MTA70, METTL3, N(6)-adenosine-methyltransferase catalytic subunit METTL3, Methyltransferase-like protein 3, N(6)-adenosine-methyltransferase 70 kDa subunit, hMETTL3, MT-A70

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: EPR18810
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

Anti-METTL3 antibody [EPR18810] (ab195352) was developed by Abcam using patented rabbit monoclonal antibody technology and is validated for use in Flow Cyt (Intra), ICC/IF, IHC-P, IP and WB.

Anti-METTL3 antibody [EPR18810] (ab195352) was first used in a scientific publication in 2016 and has been cited over 293 times in peer reviewed journals. It's performance in Western Blot and IHC in human and mouse samples is trusted by the scientific community.

Abcam's high quality manufacturing and validation processes ensure Anti-METTL3 antibody [EPR18810] (ab195352) has high sensitivity and specificity alongside high lot-to-lot consistency and reproducibility.

The specificity of Anti-METTL3 antibody [EPR18810] (ab195352) has been confirmed by Western Blot testing in METTL3 knockout Mouse embryonic stem cells cells.

Anti-METTL3 antibody [EPR18810] (ab195352) has 8 independent reviews from customers.

Anti-METTL3 antibody [EPR18810] (ab195352) specifically detects METTL3 (UniProt ID: Q86U44; Molecular weight: 65kDa) and is sold in 100 µL and 1 mL selling sizes.

Conjugation-ready, carrier free format available for antibody clone EPR18810 - Anti-METTL3 antibody [EPR18810] - BSA and Azide free ab221795.

Antibody clone EPR18810 is also available pre-conjugated to a variety of labels for your convenience - Alexa Fluor^® 488, Alexa Fluor^® 647 (Alexa Fluor® 488 Anti-METTL3 antibody [EPR18810] ab216936, Alexa Fluor® 647 Anti-METTL3 antibody [EPR18810] ab217109).

Highly-cited antibody with >400 citations. A highly specific KO validated METTL3 antibody, essential for studying RNA methylation and mRNA splicing. This antibody is crucial in epigenetic regulation research, particularly in understanding METTL3's role in gene expression and cancer progression. It is widely used in studies of METTL3 inhibitors and the METTL3-METTL14 complex. Elevated METTL3 expression is linked to cancer progression, particularly in lung adenocarcinoma, while WTAP overexpression is associated with increased cell migration and invasion in various cancers.

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.

Supplementary info

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

Activity summary

The METTL3 protein also known as methyltransferase like 3 plays an important role in the modification of RNA. It is a component of the N6-adenosine-methyltransferase complex and is responsible for introducing the m6A mark on RNA. The molecular weight of METTL3 is approximately 70 kDa. This protein shows high expression in the human central nervous system and various cancer tissues indicating its broad significance.

Biological function summary

METTL3 contributes to the post-transcriptional regulation of gene expression by modulating m6A methylation. It functions as part of a complex with METTL14 and Wilms' tumor 1-associating protein (WTAP) to execute this modification. The m6A mark influences RNA stability splicing export and translation affecting cellular processes significantly.

Pathways

METTL3 operates within critical pathways such as the mRNA stability and processing pathway. In these pathways METTL3 interacts with proteins like METTL14 and YTH domain-containing proteins which are important for m6A reader function. Together they regulate RNA processing events that influence cellular function and adaptation.

Associated diseases and disorders

METTL3 associates strongly with cancer development and neurological disorders. In cancers alterations in METTL3 expression affect tumor progression and metastasis impacting oncogenes like MYC. In the realm of neurological disorders METTL3's role in m6A modification affects RNA regulation aligning it with proteins like FTO which also participate in m6A demethylation processes. Understanding these connections highlights METTL3 as a potential target for therapeutic interventions.

Product promise

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19 product images

Immunohistochemistry - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of formalin fixed paraffin embedded human ovarian cancer labelling METTL3 with ab195352 at a concentration of 1.0µ/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32mins.
ab195352 anti-METTL3 antibody [EPR18810] was incubated for 16mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.
Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)
This image is courtesy of a customer review by Pedro Batista.
Western blot - Anti-METTL3 antibody [EPR18810] (ab195352)
Blocking step: 5% Milk for 30 minutes at 25^oC.
All lanes: Western blot - Anti-METTL3 antibody [EPR18810] (ab195352) at 1/1000 dilution
Lane 1: Mouse embryonic stem cells at 15 µg
Lane 2: Mouse embryonic stem cells (METTL3 Knock-out) at 15 µg
Secondary
All lanes: Goat anti-Rabbit Polyclonal at 1/10000 dilution
Performed under reducing conditions.
Predicted band size: 64 kDa
Exposure time: 5min
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded Rat skin tissue labeling METTL3 using ab195352 at 1/2000 dilution, followed by a Ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Counterstain: Hematoxylin. Nuclear staining on Rat skin. The section was incubated with ab195352 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Perform Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

Inset image: negative control - secondary antibody only.
Western blot - Anti-METTL3 antibody [EPR18810] (ab195352)
METTL3 Western blot staining using rabbit Anti-METTL3 antibody
In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining.
All lanes: Western blot - Anti-METTL3 antibody [EPR18810] (ab195352) at 1/1000 dilution
Lane 1: Mouse brain lysate prepared in 1%SDS Hot lysis method at 20 µg
Lane 2: Mouse brain lysate prepared in RIPA lysis method at 20 mg/mL
Lane 3: Mouse heart lysate prepared in 1%SDS Hot lysis method at 20 µg
Lane 4: Mouse heart lysate prepared in RIPA lysis method at 20 mg/mL
Lane 5: Mouse kidney lysate prepared in RIPA lysis method at 20 µg
Lanes 6 - 7: Mouse spleen lysate prepared in spleen lysis method at 20 µg
Lane 8: Rat brain lysate prepared in RIPA lysis method at 20 µg
Lane 9: Rat kidney lysate prepared in RIPA lysis method at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 64 kDa
Observed band size: 64 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded Rat testis tissue labeling METTL3 using ab195352 at 1/2000 dilution, followed by a Ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Counterstain: Hematoxylin. Nuclear staining on Rat testis. The section was incubated with ab195352 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Perform Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

Inset image: negative control - secondary antibody only.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded Mouse skin labeling METTL3 using ab195352 at 1/2000 dilution, followed by a Ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Counterstain: Hematoxylin. Nuclear staining on Mouse skin. The section was incubated with ab195352 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Perform Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

Inset image: negative control - secondary antibody only.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling METTL3 using ab195352 at 1/2000 dilution, followed by a Ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Counterstain: Hematoxylin. Nuclear staining on Mouse testis. The section was incubated with ab195352 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Perform Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

Inset image: negative control - secondary antibody only.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded Human ovarian cancer tissue labeling METTL3 using ab195352 at 1/2000 dilution, followed by a Ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Counterstain: Hematoxylin. Nuclear staining on human ovarian cancer. The section was incubated with ab195352 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Perform Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

Inset image: negative control - secondary antibody only.
Immunocytochemistry/ Immunofluorescence - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labeling METTL3 with ab195352 at 1/200 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining in NIH/3T3 cells. The nuclear counter stain is DAPI (blue). Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 was used at 1/200 dilution as the counterstain.
Immunocytochemistry/ Immunofluorescence - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HCT116 (Human colorectal carcinoma cell line) cells labeling METTL3 with ab195352 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HCT116 cell line. The nuclear counter stain is DAPI (blue).
Tubulin is detected with anti-alpha Tubulin mouse mAb (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (AlexaFluor®594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows:
1. ab195352 at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (AlexaFluor®594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) secondary antibody at 1/1000 dilution.
2. anti-alpha Tubulin mouse mAb (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded rat testis tissue labeling METTL3 using ab195352 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) secondary antibody at 1/500 dilution. Counterstain: Hematoxylin.
Inset image: negative control obtained using PBS instead of ab195352, and secondary antibody only.
Note: Nuclear staining on rat testis was observed.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Flow Cytometry (Intracellular) - Anti-METTL3 antibody [EPR18810] (ab195352)
ab195352 staining METTL3 in the human cell line HEK-293 (Human epithelial cell line from embryonic kidney) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde and the sample was incubated with the primary antibody at a 1/70 dilution, followed by Goat-Anti Rabbit IgG (Alexa Fluor^® 488) secondary antibodyat a1/2000 dilution.
Isoytype control: Rabbit monoclonal IgG (Black)
Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)
Immunoprecipitation - Anti-METTL3 antibody [EPR18810] (ab195352)
METTL3 was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab195352 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab195352 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366), was used for detection at 1/10000 dilution.
Lane 1: Input: 10μg of HeLa whole cell lysate.
Lane 2: HeLa whole cell lysate following IP with ab195352.
Lane 3: Negative control: IP using Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab195352 in HeLa whole cell lysates.
Blocking and dilution buffer and concentration: 5% NFDM/TBST. 1 second exposure.
All lanes: Immunoprecipitation - Anti-METTL3 antibody [EPR18810] (ab195352)
Predicted band size: 64 kDa
Observed band size: 67 kDa
Western blot - Anti-METTL3 antibody [EPR18810] (ab195352)
5% NFDM/TBST: Blocking and diluting buffer.
All lanes: Western blot - Anti-METTL3 antibody [EPR18810] (ab195352) at 1/5000 dilution
Lane 1: Raji (Human Burkitt's lymphoma cell line) whole cell lysate at 20 µg
Lane 2: HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 3: HEK-293 (Human epithelial cells from embryonic kidney) whole cell lysate at 20 µg
Lane 4: Jurkat (Human T cell leukemia cells from peripheral blood) whole cell lysate at 20 µg
Lane 5: NCCIT (human pluripotent embryonal carcinoma) whole cell lysate at 20 µg
Lane 6: F9 (Mouse embyro testicular cancer cell line) whole cell lysate at 20 µg
Lane 7: Neuro-2a (Mouse neuroblastoma cells) whole cell lysate at 20 µg
Lane 8: LLC1 (Mouse lung carcinoma) whole cell lysate at 20 µg
Secondary
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 64 kDa
Observed band size: 64 kDa
Exposure time: 10s
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded human bladder cancer tissue labeling METTL3 using ab195352 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) secondary antibody at 1/500 dilution. Counterstain: Hematoxylin.
Inset image: negative control obtained using PBS instead of ab195352, and secondary antibody only.
Note: Nuclear staining on human bladder cancer was observed.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling METTL3 with ab195352 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cell line. The nuclear counter stain is DAPI (blue).
Tubulin is detected with Anti-alpha Tubulin mouse MAb (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows:
1. ab195352 at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) secondary antibody at 1/1000 dilution.
2. Anti-alpha Tubulin mouse MAb (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/1000 dilution.
Western blot - Anti-METTL3 antibody [EPR18810] (ab195352)
5% NFDM/TBST: Blocking and diluting buffer.
All lanes: Western blot - Anti-METTL3 antibody [EPR18810] (ab195352) at 1/1000 dilution
Lane 1: C6 (Rat glial tumor cells) whole cell lysate at 10 µg
Lane 2: RAW 264.7 (Mouse macrophage cells transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg
Lane 3: PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate at 10 µg
Lane 4: NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate at 10 µg
Secondary
All lanes: Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/100000 dilution
Predicted band size: 64 kDa
Observed band size: 64 kDa
Exposure time: 2s
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-METTL3 antibody [EPR18810] (ab195352)
Immunohistochemical analysis of paraffin-embedded mouse testis tissue labeling METTL3 using ab195352 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) secondary antibody at 1/500 dilution. Counterstain: Hematoxylin.
Inset image: negative control obtained using PBS instead of ab195352, and secondary antibody only.
Note: Nuclear staining on mouse testis was observed.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-METTL3 antibody [EPR18810] (ab195352)
5% NFDM/TBST: Blocking and diluting buffer.
All lanes: Western blot - Anti-METTL3 antibody [EPR18810] (ab195352) at 1/5000 dilution
All lanes: Human thymus lysate at 10 µg
Secondary
All lanes: Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution
Predicted band size: 64 kDa
Observed band size: 64 kDa
Exposure time: 3min