Key features and details
- Rabbit polyclonal to CLLD8/SETDB2
- Suitable for: WB, ICC/IF
- Reacts with: Human
- Isotype: IgG
参阅全部 CLLD8/SETDB2 一抗
经测试应用适用于: WB, ICC/IFmore details
常规说明Our collaborator on this project has found that the antibody works poorly in WB.
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In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.
We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.
Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.
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存放说明Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
Preservative: 0.02% Sodium azide
Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
Concentration information loading...
纯度Immunogen affinity purified
Our Abpromise guarantee covers the use of ab5517 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use at an assay dependent concentration. Predicted molecular weight: 82 kDa.|
|ICC/IF||Use at an assay dependent concentration.|
功能Histone methyltransferase involved in left-right axis specification in early development and mitosis. Specifically trimethylates 'Lys-9' of histone H3 (H3K9me3). H3K9me3 is a specific tag for epigenetic transcriptional repression that recruits HP1 (CBX1, CBX3 and/or CBX5) proteins to methylated histones. Contributes to H3K9me3 in both the interspersed repetitive elements and centromere-associated repeats. Plays a role in chromosome condensation and segregation during mitosis.
组织特异性Ubiquitous. Highest expression in heart, testis and ovary.
序列相似性Belongs to the histone-lysine methyltransferase family.
Contains 1 MBD (methyl-CpG-binding) domain.
Contains 1 pre-SET domain.
Contains 1 SET domain.
- Information by UniProt
- C13orf4 antibody
- Chromosome 13 open reading frame 4 antibody
- Chronic lymphocytic leukemia deletion region 8 antibody
Anti-CLLD8/SETDB2 antibody (ab5517) at 1 µg/ml +
Recombinant Human CLLD8/SETDB2 protein (ab169554) at 0.01 µg
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 82 kDa
Additional bands at: 120 kDa (possible tagged protein)
Exposure time: 10 seconds
ab5517 recognises a band corresponding to the tagged CLLD8/SETB2 at approximately 120 kDa.
Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.
ICC/IF image of ab5517 stained MCF-7 cells. The cells were 4% formaldehyde 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 ab5517 at 5ug/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 Goat anti Rabbit (ab96899) IgG (H+L) used at a 1/250 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.
Left: Endogenous CLLD8
Detection using indirect fluorescence of the signal corresponding to endogenous CLLD8 in 293T cells. Cells fixed using 4% formaldehyde, blocked with PBS containing 3% milk and 0.5% Triton X-100, incubated for 1 hour at 37 °C using a 1/50 dilution of antibody ab5517. Cells were then washed 3 times and incubated for 1 hour at 37 °C with a goat anti-rabbit secondary antibody (1/500 dilution) coupled to Alexa Fluor 555 (Molecular Probes). Following 3 more washes, cells were stained with DAPI and mounted with Vectashield.
Right: Overexpressed CLLD8
Detection using indirect fluorescence of the signal corresponding to staining with anti-FLAG mouse antibody (top) and an antibody (ab5517) against CLLD8 (middle) in 293T cells. 293T cells were transfected with vectors for overexpression of flagged CLLD8 using Lipofectamine 2000 transfection procedure (Invitrogen). Cells were fixed 48 hours post-tra
Detection using indirect fluorescence of the signal corresponding to endogenous CLLD8 in 293T cells. Using Lipofectamine 2000 (Invitrogen), cells were transfected with either a control shRNA directed against luciferase (left, SiRluc) or a specific siRNA directed against CLLD8 (right). 48 hours post-transfection, cells were fixed, blocked, and incubated with a 1:50 dilution of ab5517 at 37 °C for 1 hour. Cells were then washed 3 times and incubated at 37 °C for 1 hour with a 1/500 dilution of goat anti-rabbit antibody coupled with Alexa Fluor 555 (Molecular Probes). Following 3 further washes cells were stained with DAPI and mounted with Vectashield.
Antibody signals were extinguished when a specific RNAi was used. The few cells which retained a signal were presumably not transfected.
All lanes : Anti-CLLD8/SETDB2 antibody (ab5517) at 1 µg/ml
Lane 1 : Sf21 cells infected by baculovirus containing FLAG-CLLD8 gene at 10 µg
Lane 2 : Sf21 cells infected by baculovirus containing FLAG-CLLD8 gene at 20 µg
Lane 3 : non-infected Sf21 cells at 10 µg
Lane 4 : non-infected Sf21 cells at 20 µg
Lane 5 : Sf21 infected by baculovirus containing FLAG-CLLD8 gene at 10 µg with Human CLLD8/SETDB2 peptide (ab24398) at 1 µg/ml
Lane 6 : Sf21 infected by baculovirus containing FLAG-CLLD8 gene at 20 µg with Human CLLD8/SETDB2 peptide (ab24398) at 1 µg/ml
Lane 7 : non-infected Sf21 cells at 10 µg with Human CLLD8/SETDB2 peptide (ab24398) at 1 µg/ml
Lane 8 : non-infected Sf21 cells at 20 µg with Human CLLD8/SETDB2 peptide (ab24398) at 1 µg/ml
Predicted band size: 82 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?
Additional bands at: 22 kDa (possible non-specific binding), 90 kDa (possible non-specific binding)
ab5517 recognises a band corresponding to FLAG-tagged CLLD8/SETB2 at approximately 110 kDa in Sf21 cells infected by baculovirus expressing CLLD8/SETDB2 (lanes1-2). This is larger than the predicated band size (81 kDa) due to the addition of the FLAG tag.
ab5517 does not detect a band in uninfected sf21 cells (lanes3-4) or following blocking studies using the immunizing peptide (lanes5-6).
There appears to be a ladder of proteins recognised by ab5517 (lanes1-2), which could be attributed to degradation products.
ab5517 被引用在 3 文献中.
- Nishikawaji T et al. Oncogenic roles of the SETDB2 histone methyltransferase in gastric cancer. Oncotarget 7:67251-67265 (2016). IHC . PubMed: 27572307
- Hogarth CA et al. Identification and expression of potential regulators of the mammalian mitotic-to-meiotic transition. Biol Reprod 84:34-42 (2011). WB, IHC-P ; Mouse . PubMed: 20826732
- Falandry C et al. CLLD8/KMT1F is a lysine methyltransferase that is important for chromosome segregation. J Biol Chem 285:20234-41 (2010). WB, ICC/IF ; Human . PubMed: 20404330