Immunofluorescence analysis of Metadherin (C-Term) was done on 70% confluent log phase U-2 OS cells. The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton™ X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with ABfinity™ Metadherin (C-Term) Recombinant Rabbit Monoclonal Antibody (701171) at a dilution of 1:400 in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Flour® 488 Goat Anti-Rabbit IgG Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor 594 Phalloidin (A12381). Panel d is a merged image showing cytoplasmic localization of Metadherin (C-Term). Panel e shows no primary antibody. The images were captured at 20X magnification.
|Tested species reactivity||Human, Mouse|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Peptide corresponding to amino acids 561–570 of human Metadherin (C-term)|
|Contains||0.09% sodium azide|
|Storage Conditions||Maintain refrigerated at 2-8°C for up to 1 month. For long term storage store at -20°C|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Immunocytochemistry (ICC)||1-3 ug/ml|
|Immunohistochemistry (Paraffin) (IHC (P))||1:10-1:100|
|Western Blot (WB)||1-3 ug/ml|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
This antibody is predicted to react with non-human primate, mouse and rat based on sequence homology.
ABfinity™ recombinant antibodies are rabbit monoclonal antibodies, unmatched for producing superior results. ABfinity™ antibodies are developed by immunizing animals, screening for functionality, cloning the immunogen-specific antibody genes into high-level mammalian expression vectors, produced on a large scale, and purified with Protein A.
ABfinity™ monoclonal antibodies resemble rabbit monoclonals isolated from serum or produced by hybridomas, but demonstrate greater specificity and sensitivity. Because ABfinity™ recombinant antibodies are derived from cloned DNA sequences of the heavy and light antibody chains, they are not susceptible to cell-line drift or lot-to-lot variation, thus allowing for peak specificity and performance.
Intact IgG appears on a non-reducing gel as ~150 kDa band and upon reduction generating a ~25 kDa light chain band and a ~50 kDa heavy chain.
Metadherin (MTDH) was originally identified as an oncogene, and plays a role in promoting proliferation, invasion, survival, and enhancing chemoresistance. Expression of MTDH is associated with Ha-ras, PI3K/Akt, NF- kappaB and Wnt/ beta-catenin signaling pathways. Immunoflurescence and histochemical analysis has shown perinuclear, nucleolar, and cytoplasmic staining, while upregulated MTDH expression has been shown to enhance nuclear localization. The protein has three putative lysine-rich nuclear localization signals and this antibody has been raised specifically against the NLS3 region.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.