In Western blot, this antibody gave a positive signal in chicken liver tissue lysate as well as in the following whole cell lysates: HeLa; Jurkat; A431; HEK293; MCF7; U20S. This antibody also gave a positive signal in HeLa whole cell lysates in Immunofluorescence and Human Testis tissue lysate in Immunohistochemistry.
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.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Detects a band of approximately 120 kDa (predicted molecular weight: 114 kDa).Can be blocked with Human RNF20 peptide (ab32628).
Use a concentration of 1 µg/ml.
1/50. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use at an assay dependent concentration.
Component of the RNF20/40 E3 ubiquitin-protein ligase complex that mediates monoubiquitination of 'Lys-120' of histone H2B (H2BK120ub1). H2BK120ub1 gives a specific tag for epigenetic transcriptional activation and is also prerequisite for histone H3 'Lys-4' and 'Lys-79' methylation (H3K4me and H3K79me, respectively). It thereby plays a central role inb histone code and gene regulation. The RNF20/40 complex forms a H2B ubiquitin ligase complex in cooperation with the E2 enzyme UBE2A or UBE2B; reports about the cooperation with UBE2E1/UBCH are contradictory. Required for transcriptional activation of Hox genes. Recruited to the MDM2 promoter, probably by being recruited by p53/TP53, and thereby acts as a transcriptional coactivator. Mediates the polyubiquitination of isoform 2 of PA2G4 in cancer cells leading to its proteasome-mediated degradation.
Expressed in the normal brain and also in malignant gliomas (at protein level).
Protein modification; protein ubiquitination.
Belongs to the BRE1 family. Contains 1 RING-type zinc finger.
ab32629 staining RNF20 in human testis, showing a distinct and strong staining pattern in ductus seminiferus cells. Paraffin embedded human skin tissue was incubated with ab32629 (1/50 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6.
ab32629 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org
Immunocytochemistry/ Immunofluorescence - Anti-RNF20 antibody (ab32629)This image is courtesy of an Abreview submitted by Dr Kirk McManus
ab32629 (1/500) staining RNF20 in assynchronous HeLa cells (green). Cells were fixed in paraformaldehyde, permeabilised with 0.5% Triton X100/PBS and counterstained with DAPI to highlight the nucleus (red). Please refer to abreview for further experimental details.
ICC/IF image of ab32629 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32629, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) Hek293, HepG2 and MCF7 cells at 1µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 1µg/ml.
Western blot - Anti-RNF20 antibody (ab32629)Image courtesy of an anonymous Abreview.
All lanes : Anti-RNF20 antibody (ab32629) at 1/1000 dilution
Lane 1 : Whole cell lysate prepared from Ntera2 cells Lane 2 : Whole cell lysate prepared from Ntera2 cells + siRNA against RNF20 for 48 hours
Lysates/proteins at 20 µg per lane.
Secondary All lanes : HRP conjugated goat polyclonal at 1/10000 dilution