Recombinant Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855]

Product name

Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free
See all RNA polymerase II CTD repeat YSPTSPS primary antibodies
Description

Rabbit monoclonal [EPR18855] to RNA polymerase II CTD repeat YSPTSPS (phospho S2) - BSA and Azide free
Host species

Rabbit
Tested applications

Suitable for: Flow Cyt (Intra), WB, Dot blot, ICC/IF, IP, IHC-P, ChIPmore details
Species reactivity

Reacts with: Mouse, Rat, Human
Predicted to work with: Saccharomyces cerevisiae
Immunogen

Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
Positive control
- IHC-P: Mouse kidney tissue. ChIP: HeLa cells.
General notes
ab238450 is the carrier-free version of ab193468.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

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

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C. Do Not Freeze.
Storage buffer

pH: 7.2
Constituent: PBS
Carrier free

Yes
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR18855
Isotype

IgG
Research areas

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab238450 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application	Abreviews	Notes
Flow Cyt (Intra)		Use at an assay dependent concentration.
WB		Use at an assay dependent concentration. Detects a band of approximately 270 kDa (predicted molecular weight: 192 kDa).
Dot blot		Use at an assay dependent concentration.
ICC/IF		Use at an assay dependent concentration.
IP		Use at an assay dependent concentration.
IHC-P		Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. Recommended for mouse only.
ChIP		Use at an assay dependent concentration.

Notes
Flow Cyt (Intra) Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 270 kDa (predicted molecular weight: 192 kDa).
Dot blot Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. Recommended for mouse only.
ChIP Use at an assay dependent concentration.

Function

DNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates. Largest and catalytic component of RNA polymerase II which synthesizes mRNA precursors and many functional non-coding RNAs. Forms the polymerase active center together with the second largest subunit. Pol II is the central component of the basal RNA polymerase II transcription machinery. It is composed of mobile elements that move relative to each other. RPB1 is part of the core element with the central large cleft, the clamp element that moves to open and close the cleft and the jaws that are thought to grab the incoming DNA template. At the start of transcription, a single-stranded DNA template strand of the promoter is positioned within the central active site cleft of Pol II. A bridging helix emanates from RPB1 and crosses the cleft near the catalytic site and is thought to promote translocation of Pol II by acting as a ratchet that moves the RNA-DNA hybrid through the active site by switching from straight to bent conformations at each step of nucleotide addition. During transcription elongation, Pol II moves on the template as the transcript elongates. Elongation is influenced by the phosphorylation status of the C-terminal domain (CTD) of Pol II largest subunit (RPB1), which serves as a platform for assembly of factors that regulate transcription initiation, elongation, termination and mRNA processing. Acts as an RNA-dependent RNA polymerase when associated with small delta antigen of Hepatitis delta virus, acting both as a replicate and transcriptase for the viral RNA circular genome.
Sequence similarities

Belongs to the RNA polymerase beta' chain family.
Domain

The C-terminal domain (CTD) serves as a platform for assembly of factors that regulate transcription initiation, elongation, termination and mRNA processing.
Post-translational
modifications

The tandem heptapeptide repeats in the C-terminal domain (CTD) can be highly phosphorylated. The phosphorylation activates Pol II. Phosphorylation occurs mainly at residues 'Ser-2' and 'Ser-5' of the heptapeptide repeat and is mediated, at least, by CDK7 and CDK9. CDK7 phosphorylation of POLR2A associated with DNA promotes transcription initiation by triggering dissociation from DNA. Phosphorylation also takes place at 'Ser-7' of the heptapeptide repeat, which is required for efficient transcription of snRNA genes and processing of the transcripts. The phosphorylation state is believed to result from the balanced action of site-specific CTD kinases and phosphatases, and a 'CTD code' that specifies the position of Pol II within the transcription cycle has been proposed. Dephosphorylated by the protein phosphatase CTDSP1.
Among tandem heptapeptide repeats of the C-terminal domain (CTD) some do not match the Y-S-P-T-S-P-S consensus, the seventh serine residue 'Ser-7' being replaced by a lysine. 'Lys-7' in these non-consensus heptapeptide repeats can be alternatively acetylated, methylated and dimethylated. EP300 is one of the enzyme able to acetylate 'Lys-7'. Acetylation at 'Lys-7' of non-consensus heptapeptide repeats is associated with 'Ser-2' phosphorylation and active transcription. It may regulate initiation or early elongation steps of transcription specially for inducible genes.
Methylated at Arg-1810 prior to transcription initiation when the CTD is hypophosphorylated, phosphorylation at Ser-1805 and Ser-1808 preventing this methylation. Symmetrically or asymmetrically dimethylated at Arg-1810 by PRMT5 and CARM1 respectively. Symmetric or asymmetric dimethylation modulates interactions with CTD-binding proteins like SMN1/SMN2 and TDRD3. SMN1/SMN2 interacts preferentially with the symmetrically dimethylated form while TDRD3 interacts with the asymmetric form. Through the recruitment of SMN1/SMN2, symmetric dimethylation is required for resolving RNA-DNA hybrids created by RNA polymerase II, that form R-loop in transcription terminal regions, an important step in proper transcription termination. CTD dimethylation may also facilitate the expression of select RNAs. Among tandem heptapeptide repeats of the C-terminal domain (CTD) some do not match the Y-S-P-T-S-P-S consensus, the seventh serine residue 'Ser-7' being replaced by a lysine. 'Lys-7' in these non-consensus heptapeptide repeats can be alternatively acetylated, methylated and dimethylated. Methylation occurs in the earliest transcription stages and precedes or is concomitant to 'Ser-5' and 'Ser-7' phosphorylation.
Ubiquitinated by WWP2 leading to proteasomal degradation (By similarity). Following UV treatment, the elongating form of RNA polymerase II (RNA pol IIo) is ubiquitinated UV damage sites without leading to degradation: ubiquitination is facilitated by KIAA1530/UVSSA and promotes RNA pol IIo backtracking to allow access to the nucleotide excision repair machinery.
Cellular localization

Nucleus.
Target information above from: UniProt accession P24928 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 5430 Human
- Entrez Gene: 20020 Mouse
- Entrez Gene: 363633 Rat
- Entrez Gene: 851415 Saccharomyces cerevisiae
- Omim: 180660 Human
- SwissProt: P24928 Human
- SwissProt: P08775 Mouse
- SwissProt: P04050 Saccharomyces cerevisiae
- Unigene: 270017 Human
- Unigene: 16533 Mouse
see all
Alternative names
- DNA directed RNA polymerase II A antibody
- DNA-directed RNA polymerase II largest subunit RNA polymerase II 220 kd subunit antibody
- DNA-directed RNA polymerase II subunit A antibody
- DNA-directed RNA polymerase II subunit RPB1 antibody
- DNA-directed RNA polymerase III largest subunit antibody
- hRPB220 antibody
- hsRPB1 antibody
- POLR2 antibody
- Polr2a antibody
- POLRA antibody
- Polymerase (RNA) II (DNA directed) polypeptide A 220kDa antibody
- Polymerase (RNA) II (DNA directed) polypeptide A antibody
- RNA polymerase II subunit B1 antibody
- RNA-directed RNA polymerase II subunit RPB1 antibody
- RPB1 antibody
- RPB1_HUMAN antibody
- RPBh1 antibody
- RpIILS antibody
- RPO2 antibody
- RPOL2 antibody
see all

Immunoprecipitation - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

RNA polymerase II CTD repeat YSPTSPS (phospho S2) was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab193468 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab193468 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1: HeLa whole cell lysate 10µg (Input).

Lane 2: ab193468 IP in HeLa whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab193468 in HeLa whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 1 second.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
ChIP - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Chromatin was prepared from HeLa cells according to the Abcam Dual-X-ChIP protocol*. Cells were fixed with 1.5 mM EGS for 30mins and then formaldehyde for 10min.
The ChIP was performed with 25 μg of chromatin, 5 μg of ab193468 (red), or 5 μg of rabbit normal IgG ab172730 (gray) and 20 μL of Protein A/G sepharose beads. The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).
Primers and probes are located in the first kb of the transcribed region

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).

*http://www.abcam.com/resources?keywords=X%20ChIP%20protocol
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on mouse spleen is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on epithelium cells and glomerulus cells of mouse kidney is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on mouse testis is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized MCF7 (Human breast adenocarcinoma cell line) cells labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on MCF7 cell line. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab193468 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Immunocytochemistry/ Immunofluorescence - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cell line. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab193468 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Immunocytochemistry/ Immunofluorescence - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized PC-12 (Rat adrenal gland pheochromocytoma) cells labeling RNA polymerase II CTD repeat YSPTSPS (phospho S2) with ab193468 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on PC-12 cell line. The nuclear counterstain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab193468 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Flow Cytometry (Intracellular) - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling RNA polymerase II CTD repeat YSPTSPS antibody (phospho S2) (red) with purified ab193468 at a dilution of 1/70. Goat anti rabbit IgG (Alexa Fluor^® 488) was used as the secondary antibody at 1/2000. Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Isotype control antibody was Rabbit monoclonal IgG (black). The blue line shows cells without incubation with primary antibody and secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Dot Blot - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Dot blot analysis of RNA polymerase II CTD repeat YSPTSPS (phospho S2) peptide (Lane 1) and non-phospho peptide (Lane 2) labeled using ab193468 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated secondary antibody (ab97051) at 1/100000 dilution.
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Dot Blot - Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

Lane 1: RNA polymerase II CTD repeat YSPTSPS (phospho S2) phospho peptide
Lane 2: RNA polymerase II CTD repeat YSPTSPS non-phospho peptide (peptide of ab193468)
Lane 3: RNA polymerase II CTD repeat YSPTSPS (phospho S5) phospho peptide
Lane 4: RNA polymerase II CTD repeat YSPTSPS non-phospho peptide (peptide of ab193467)
Labeled using ab193468 at 1/1000 dilution (0.8 ug/ml), followed by Goat Anti-Rabbit IgG H&L (HRP) secondary antibody (ab97051) at 1/100000 dilution.
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab193468).
Anti-RNA polymerase II CTD repeat YSPTSPS (phospho S2) antibody [EPR18855] - BSA and Azide free (ab238450)

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

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Overview

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Description

Host species

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Carrier free

Purity

Clonality

Clone number

Isotype

Research areas

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Target

Function

Sequence similarities

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Post-translationalmodifications

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Post-translational
modifications