Overview

  • Product name
    Anti-RING2 / RING1B / RNF2 antibody - ChIP Grade
    See all RING2 / RING1B / RNF2 primary antibodies
  • Description
    Goat polyclonal to RING2 / RING1B / RNF2 - ChIP Grade
  • Host species
    Goat
  • Specificity
    This antibody recognises a band at just above the expected size of 37kD.
  • Tested applications
    Suitable for: ICC/IF, WB, ChIPmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide:

    EAGPSNKRTKTSDC

    , corresponding to amino acids 189-201 of Human RING1B.

  • General notes

    RING1B interacts with CP2, a mammalian member of the grainyhead-like family of transcription factors, in vitro and in vivo. The functional consequence of this interaction is repression of CP2-dependent transcription.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 0.42% Potassium phosphate, 0.87% Sodium chloride
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Primary antibody notes
    RING1B interacts with CP2, a mammalian member of the grainyhead-like family of transcription factors, in vitro and in vivo. The functional consequence of this interaction is repression of CP2-dependent transcription.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab3832 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
ICC/IF 1/300. (see Abreviews)
WB Use at an assay dependent dilution. Detects a band of approximately 43 kDa (predicted molecular weight: 37.6 kDa).
ChIP Use at an assay dependent dilution.

Target

  • Function
    E3 ubiquitin-protein ligase that mediates monoubiquitination of 'Lys-119' of histone H2A, thereby playing a central role in histone code and gene regulation. H2A 'Lys-119' ubiquitination gives a specific tag for epigenetic transcriptional repression and participates in X chromosome inactivation of female mammals. May be involved in the initiation of both imprinted and random X inactivation. Essential component of the Polycomb group (PcG) multiprotein PRC1 complex, a complex required to maintain the transcriptionally repressive state of many genes, including Hox genes, throughout development. PcG PRC1 complex act via chromatin remodeling and modification of histones, rendering chromatin heritably changed in its expressibility. E3 ubiquitin-protein ligase activity is enhanced by BMI1/PCGF4. Acts as the main E3 ubiquitin ligase on histone H2A of the PRC1 complex, while RING1 may rather act as a modulator of RNF2/RING2 activity.
  • Pathway
    Protein modification; protein ubiquitination.
  • Sequence similarities
    Contains 1 RING-type zinc finger.
  • Post-translational
    modifications
    Polyubiquitinated in the presence of UBE2D3 (in vitro).
    Monoubiquitinated, by auto-ubiquitination.
  • Cellular localization
    Nucleus. Chromosome. Enriched on inactive X chromosome (Xi) in female trophoblast stem (TS) cells as well as differentiating embryonic stem (ES) cells. The enrichment on Xi is transient during TS and ES cell differentiation. The association with Xi is mitotically stable in non-differentiated TS cells.
  • Information by UniProt
  • Database links
  • Alternative names
    • BAP 1 antibody
    • BAP1 antibody
    • DING antibody
    • DinG protein antibody
    • E3 ubiquitin protein ligase RING 2 antibody
    • E3 ubiquitin protein ligase RING2 antibody
    • E3 ubiquitin-protein ligase RING2 antibody
    • HIP2 interacting protein 3 antibody
    • HIP2-interacting protein 3 antibody
    • HIPI 3 antibody
    • HIPI3 antibody
    • Huntingtin interacting protein 2 interacting protein 3 antibody
    • Huntingtin-interacting protein 2-interacting protein 3 antibody
    • OTTHUMP00000060668 antibody
    • Polycomb M33 interacting protein Ring 1B antibody
    • Polycomb M33 interacting protein Ring1B antibody
    • Protein DinG antibody
    • RING 1B antibody
    • RING 2 antibody
    • RING finger protein 1B antibody
    • RING finger protein 2 antibody
    • RING finger protein BAP 1 antibody
    • RING finger protein BAP-1 antibody
    • RING finger protein BAP1 antibody
    • RING1b antibody
    • RING2_HUMAN antibody
    • RNF 2 antibody
    • Rnf2 antibody
    see all

Images

  • Western blot using ab3832 at 1/500. Lysates at 20µg/lane.

    Lane 1: 3T3 cell lysate
    Lane 2: U937 cell lysate
    Lane 3: Jurkat cell lysate
    Lane 4: Mouse brain lysate
    Lane 5: CHO-K1 cell lysate

    Secondary ab: rabbit polyclonal to goat IgG ab6741 1/5000
    Exposure time: 3 minutes

    Western blot using ab3832 at 1/500. Lysates at 20µg/lane.

    Lane 2: U937 cell lysate
    Lane 3: Jurkat cell lysate
    Lane 4: Mouse brain lysate
    Lane 5: CHO-K1 cell lysate

    Secondary ab: rabbit polyclonal to goat IgG ab6741 1/5000 Exposure time: 3 minutes
  • ab3832 staining RING2/RING1B/RNF2 in Human Hela cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with methanol and blocked with 0.2% fish scale gelatin for 1 hour at 25°C. Samples were incubated with primary antibody (1/300 in PBS + 0.2% fish scale gelatin) for 20 minutes at 25°C. An Alexa Fluor®488-conjugated Donkey anti-goat IgG polyclonal (1/500) was used as the secondary antibody.

    See Abreview

References

This product has been referenced in:
  • Rothberg JLM  et al. Mtf2-PRC2 control of canonical Wnt signaling is required for definitive erythropoiesis. Cell Discov 4:21 (2018). Read more (PubMed: 29736258) »
  • Pulikkan JA  et al. CBFß-SMMHC Inhibition Triggers Apoptosis by Disrupting MYC Chromatin Dynamics in Acute Myeloid Leukemia. Cell 174:172-186.e21 (2018). Read more (PubMed: 29958106) »
See all 19 Publications for this product

Customer reviews and Q&As

1-5 of 5 Abreviews or Q&A

Answer

Thank you for taking the time to contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

I would like to reassure you that this antibody is tested and covered by our 6 month guarantee forWB and in human and mouse samples. In the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund.

I would like to investigate this particular case further for you, and also obtain more information for our quality monitoring records.with furhter information, I woudl be more able to provide further suggestion onthe western blot. In order to proceed with this, I have enclosed a technical questionnaire below. I would appreciate if you are able to complete this. It will help you put the information we require together very easily.

Thank you for your time and cooperation. We look forward to receiving the completed questionaire.



Order Details
Antibody code:RING1b (ab3832) antibody

Problem
Choose: Non-specific band Multiple bands No signal or weak signal High background

Lot number

Purchase order number
or preferably Abcam order number:



General Information
Antibody storage conditions (temperature/reconstitution etc)


Description of the problem (high background, wrong band size, more bands, no band etc.)


Sample (Species/Cell extract/Nuclear extract/Purified protein/Recombinant protein etc.)


Sample preparation (Buffer/Protease inhibitors/Heating sample etc.)


Amount of protein loaded
20µg

Electrophoresis/Gel conditions (Reducing or Non-reducing gel, % of the gel etc.)


Transfer and blocking conditions (Buffer/time period, Blocking agent etc.)


Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)

1:500 in PBS-Tween for overnight incubation.

Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)


Detection method (ECL, ECLPlus etc.)

HCL exposure

Positive and negative controls used (please specify)



Optimization attempts (problem solving)
How many times have you tried the Western?



Have you run a "No Primary" control?
Yes No

Do you obtain the same results every time?
Yes No
e.g. are the background bands always in the same place?


What steps have you altered?


Additional Notes:


Image:
We would appreciate if you are able to provide an image (including molecular weight markers) which would help us to asess the results.

Read More

Answer

Thank you for contacting us.
I am sorry to hear the antibody does not give you results as expected.

I do have a couple questions in order to understand your experiment better:

1) Are you using the antibody for IP or ChIP? There are differences between both techniques including the cross-linking step and the detection method. IP by itself has not been tested with this antibody.

2) Are you using WB (for IP) to obtain your results or rather RT-PCR (for ChIP)?
3) Also, what controls have you included in your experiment, especially when doing ChIP?
4) Could you please send me your protocol?

I look forward to hear back from you.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (Hela cells)
Specification
Hela cells
Fixative
Methanol
Permeabilization
No
Blocking step
Fish scale gelatin in PBS as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 0.2% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Aug 02 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Jurkat cells)
Loading amount
100000 cells
Specification
Jurkat cells
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted May 17 2010

Answer

Thank you for your enquiry. I have consulted the EXPASY databases and I retrieved the following information: Mel18 - Tissue specificity: Detected in all tissues examined with high expression found in placenta, lung and kidney and low expression, in liver, pancreas and skeletal muscle. Ring1B - No tissue specificity information. The recommended positive control for Mel18 is human lung lysate. For Ring1B we do not have a recommended positive control. However, a reviewer has successfully used this antiserum in western blotting using mouse cultured cells (3T3) and mouse brain. This may be a useful alternative to HeLa cells as I cannot guarantee that these cells express Ring1B. I have obtained the following details of how the positive control to how Mel18 was prepared and run by western blotting. Western Blot: Approx 40 kDa band observed in Human Lung extracts (predicted MW of 40kDa according to NP_009075). Recommended for use at 1-2µg/ml. Tissue Lysis. Tissue chunks were weighed and cut into approx 1mm cubes using a razor blade. The tissue was transferred to a handheld homogenizer and 3 ml of ice-cold RIPA buffer was added per 1g of tissue. The tissue was gently homogenised over 20 minutes on ice. The resulting lysate was aliquotted into 1.5 ml microfuge tubes and centrifuged at 13,000 rpm for 5 min in a microfuge. The supernatant was transferred into clean tubes and its protein concentration was measured with BioRad protein assay. The concentration was then adjusted to 5 mg/ml with RIPA lysis buffer. An equal volume of 2 x SDS sample buffer was added and the cell lysate was boiled for 5 minutes. Lysates were stored at -80C until use.(RIPA buffer = 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM PMSF, 1 mM EDTA, 5 µg/ml Aprotinin, 5 µg/ml Leupeptin, 1% Triton X-100, 1% Sodium deoxycholate, 0.1% SDS). SDS PAGE. Samples were run at 200V constant on a 12% acrylamide SDS-PAGE mini gel - using Biorad Mini-Protean 3 kit and protocols. Before loading samples had 5% (v/v) 2-ME added and were boiled for 3 minutes. Transfer. We used a Biorad Mini Trans-Blot, constant 100 V for 1 hour. Transfer Buffer was 20 mM Tris pH 8.0, 150 mM Glycine, 10% Methanol. We transferred to Millipore PVDF membrane and stained with Ponceau Red to evaluate the transfer. Staining. The membrane was blocked in 2.5% skimmed milk in TBS-T (TBS + 0.05% Tween-20) for 1 hr at room temperature with agitation. Primary antibody was incubated for 1 hr at room temperature with agitation. We used [a competitor's] secondary 1:3000) for 1 hr at room temperature with agitation. We washed with TBST three times after primary and secondary antibody, each wash lasting for 5-10 mins. ECL-plus (Amersham) was used rather than ECL, which is considerably more sensitive. Final detection was on autoradiography film. I hope this information helps. Please do not hesitate to contact me should you require further assistance.

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Sign up