Product nameAnti-Sumo 2+3 antibody
See all Sumo 2+3 primary antibodies
DescriptionRabbit polyclonal to Sumo 2+3
SpecificityRecognises 2 bands representing Sumo 2 and Sumo 3 at 15 and 18kDa in Hela Nuclear extract by Western blotting.
Tested applicationsSuitable for: ICC/IF, WB, IHC-Pmore details
Species reactivityReacts with: Mouse, Human, Xenopus laevis, Chinese hamster
Predicted to work with: Rat, Cow, Pig, Zebrafish
- WB: HeLa Nuclear cell lysate. IHC-P: Human colon adenocarcinoma tissue. ICC-IF: HepG2 cells.
Storage instructionsShipped 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 bufferPreservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4
Concentration information loading...
PurityImmunogen affinity purified
Our Abpromise guarantee covers the use of ab3742 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/1000. Detects a band of approximately 15, 18 kDa (predicted molecular weight: 11.6 , 10.8 kDa).|
|IHC-P||1/800. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
RelevanceSUMO proteins, such as Sumo 2 and Sumo 3, post-translationally modify numerous cellular proteins and affect their metabolism and function. However, unlike ubiquitination, which targets proteins for degradation, sumoylation participates in a number of cellular processes, such as nuclear transport, transcriptional regulation, apoptosis, and protein stability. Sumo 2 and Sumo 3 are highly homologous, hence it is very difficult to produce antibodies which distinguish them.
Cellular localizationCytoplasmic (SUMO3) and Nuclear (SUMO2)
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Left-hand images show uninfected cells and the co-localization of SUMO-2/3 with PML (red) in control (upper rows of each block of 4 images) and PML depleted (low rows of each block of 4 images) HepaRG cells. Right-hand images show typical examples of recruitment of the indicated proteins to sites associated with HSV-1 genomes (ICP4; red) in cells at the edges of ICP0 null mutant (ΔICP0) plaques in control and PML depleted HepaRG cells. Scale bars indicate 5 µm.
Cells on glass coverslips were fixed with 1.5% formaldehyde in PBS containing 2% sucrose then treated with 0.5% Nonidet P40 substitute in PBS/10% sucrose. SUMO-2/3 was detected with ab3742. An Alexa-conjugated anti-rabbit IgG was used as the secondary antibody.
Lanes 1 & 3: Sumo 2+3 antibody (ab3742) at 1/500 dilution
Lanes 2 & 4: Sumo 2+3 antibody (ab3742) at 1/1000 dilution
Lanes 1-4: HeLa nuclear extract at 20 ug
Goat polyclonal to Rabbit IgG H&L (HRP) (ab6721) at 1/2000 dilution developed using the ECL technique
Performed under reducing conditions.
Exposure time: 1 minute
Predicted band sizes : 11.6 & 10.
ab3742 stained in HepG2 cells. Cells were fixed with 100% methanol (5min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab3742 at 5µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilution overnight at +4°C. The secondary antibodies were ab150120 (pseudo-colored red) and ab150081 (colored green) used at 1 ug/ml for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43µM for 1hour at room temperature
IHC image of Sumo 2+3 staining in human colon adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab3742, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Image courtesy of Human Protein Atlas
ab3742 staining Sumo 2 + 3 in Human skin. The paraffin embedded human skin tissue was incubated with ab3742 (1/800 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6. Ab3742 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
Immunofluorescent imaging of human cells (U2OS) with ab3742 confirms the specificity of this antibody. Antibody signal is localised exclusively to the nucleus, with diffuse background staining of the nucleoplasm. Intense foci of staining are also evident, corresponding to SUMO-2/3 accumulation in nuclear subdomains such as the PML body. This image is in exact agreement with several published reports (see for example Saitoh H et al.).
IF was performed with a standard paraformaldehyde technique (fixed in PBS buffered PFH 4% for 5 minutes, permeabilised with 0.5% triton-PBS for 5 minutes, blocked with 5% milk / 0.2% tween for one hour. Primary antibody used at 1/100 in 5% milk / 0.2% TWEEN for one hour, secondary antibody for 30 minutes. All blocking and incubation steps carried out at 37 degrees. Nuclei are stained with Hoechst stain.
ab3742 staining Sumo 2+3 in Xenopus laevis oocyte cell by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with NP-40 0.1% in PBS and blocked with 3% BSA for 60 minutes at 23°C. Samples were incubated with primary antibody (1/250 in PBS + 3% BSA) for 1 hour at 23°C. An undiluted Alexa Fluor®488-conjugated Goat anti-rabbit polyclonal was used as the secondary antibody.
This product has been referenced in:
- Chowdhury D et al. p38 MAPK pathway-dependent SUMOylation of Elk-1 and phosphorylation of PIAS2 correlate with the downregulation of Elk-1 activity in heat-stressed HeLa cells. Cell Stress Chaperones 24:393-407 (2019). Read more (PubMed: 30783905) »
- Rousseaux MW et al. Depleting Trim28 in adult mice is well tolerated and reduces levels of a-synuclein and tau. Elife 7:N/A (2018). Read more (PubMed: 29863470) »