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Also for UBA1 (NM_003334)
|Anti-Ubiquitin Activating Enzyme E1 antibody was prepared from whole rabbit serum produced by repeated immunizations with a recombinant protein corresponding to full length Human Ubiquitin Activating Enzyme E1.|
||ELISA: 1:2,000 - 1:10,000, WB: 1:1,000 - 1:5,000, IHC: 2 mg/ml - 20 ug/ml
|0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2|
|Ubiquitin Activating Enzyme (E1), also known as A1S9 and UBE1, is responsible for the first step in ubiquitin-protein isopeptide bond formation. E1 catalyzes the activation of the C-terminal carboxyl group of ubiquitin by forming a high-energy thioester bond in an ATP-dependent manner. UBE1 is monomeric and contains two active sites within the E1 molecule, allowing it to bind two ubiquitin moieties at a time, with a new ubiquitin forming an adenylate intermediate as the previous one is transferred to the thiol site. Alternative splicing results in 2 transcript variants encoding the same protein, but with different 5' UTR. Isoform 1 has a different 5' noncoding exon compared to isoform 2. Both variants encode the same protein.
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Western blot using purified anti-Ubiquitin Activating Enzyme (E1) antibody shows detection of a band at ~118 kDa corres-ponding to UBE1 (lane 1 800 nm channel). Approximately 35 µg of an A431 whole cell lysate was separated on a 4-20% Tris-Glycine gel by SDS-PAGE and transferred onto nitrocellulose. After blocking the membrane was probed with the primary antibody diluted to 1:1,000. Incubation was for 2 h at room temperature followed by washes and reaction with a 1:10,000 dilution of IRDye™800 conjugated Gt-a-Rabbit IgG [H&L] MX10 (611-132-122) for 45 min at room temperature. Molecular weight markers are shown in lane 2 (700 nm channel). IRDye™800 fluorescence image was captured using the Odyssey® Infrared Imaging System developed by LI-COR. IRDye is a trademark of LI-COR, Inc. Other detection systems will yield similar results.
Affinity Purified anti-Ubiquitin Activating Enzyme antibody was used at a 10 µg/ml to detect UBE1 in a variety of tissues including adrenal, breast, colon (epithelium), kidney, liver, lung (respiratory epithelium), ovary (oocyte and endothelium), pancreas (islet and exocrine), placenta, prostate (epithelium), skin (epithelium), spleen (lymphocytes), stomach (chief), testis, thymus, tonsil, and uterus (glandular, stroma). In many cells a punctate nuclear staining was observed. Other cells showed both cytoplasmic and nuclear staining. This image shows UBE1 staining of human lung tissue. Tissue was formalin-fixed and paraffin embedded. Personal Communi-cation, Tina Roush, LifeSpanBiosciences, Seattle, WA.