Host/IsotypeMouse / IgG2b
Clone27B10.4
Species ReactivityAll species
Validation DataWhite PaperCustomer Testmimonials
Anti-phosphotyrosine antibody is a tyrosine phosphorylation specific antibody that is part of the Signal-Seeker™ product line. APY03 anti-phosphotyrosine mouse monoclonal antibody clone 27B10 has been tested in western blot, immunofluorescence, immunoprecipitation and ELISA with excellent specificity and low background with strong potent signals. APY03 phosphotyrosine antibody recognizes a wide range of tyrosine phosphorylated proteins in cells treated with kinase inhibitors, hydrogen peroxide (H2O2) and sodium vanadate (Figure 1).
APY03 is a mouse monoclonal antibody 27B10 that recognizes phosphorytated tyrosine residues. APY03 was raised against a proprietary mixture of phosphotyrosine peptides conjugated to KLH. APY03 can detect 10 ng of phosphotyrosine-labeled bovine serum albumin (see Certificate of Analysis [COA]). APY03 is purified by protein G affinity chromatography and is supplied as a lyophilized white powder. Each Lot of antibody is quality controlled to provide a high batch to batch consistency. The Lot specific µg per tube can be found in the Lot specific COA documents at www.cytoskeleton.com. APY03 shows high specificity to phosphotyrosine peptides and does not cross-react with phosphoserine/threonine peptides in a western blot or an ELISA assay (Figures 1 & 4).
Figure 1: Western Blot using Phosphotyrosine Antibody
APY03 is highly specific for p-Tyr and does not recognize p-Ser or p-Thr tagged proteins. To see the full Western blot protocol, see the product datasheet.
Figure 2: Immunoprecipitation using Phosphotyrosine Antibody
APY03 was able to enrich a wide range of tyrosine-phosphorylated proteins from NIH3T3 cells treated with H2O2-activated orthovanadate. No signal was detected with Protein G bead control without APY03. To see the full Immunoprecipitation protocol, see the product datasheet.
Human epidermoid carcinoma A431 cells, untreated (3A) or treated (3B) with EGF (100 ng/ml for 3 minutes), and NIH3T3, untreated (3C) or treated (3D) with H2O2-activated sodium orthovanadate (100 µM for 10 minutes), were stained as described in the method. Phosphotyrosine and nuclei were visualized in green fluorescence and blue DAPI staining, respectively. To see the full Immunofluorescence protocol, see the product datasheet.
Figure 4: ELISA using Phosphotyrosine Antibody
Data show that APY03 does not crossreact with phosphothreonine and phoophoserine modified BSA. To see the full ELISA protocol, see the product datasheet.
For more information contact: signalseeker@cytoskeleton.com
Associated Products:
Signal-Seeker™ Phosphotyrosine Detection Kit (Cat. # BK160)
Signal-Seeker™ Phosphotyrosine Affinity Beads (Cat.# APY03-beads)
Signal-Seeker™: BlastR™ Rapid Lysate Prep Kit (Cat. # BLR01)
For product Datasheets and MSDSs please click on the PDF links below.
Sample Size Datasheet (Cat. APY03-S): 
Certificate of Analysis: Lot 011
For the most recent publications citing this and other Signal-Seeker™ products, see our Signal-Seeker™ Validation Data Page click here Customer Testimonial & Results Reviewed Product: Anti-Phosphotyrosine Mouse Monoclonal Antibody (Cat. # APY03) Application:Western blot “One of our lab’s primary focus is examining the signaling cascade of ouabain on the sodium pump, atp1a4. We examine the broad effects of this cascade by looking at the phosphorylation patterns of proteins, specifically tyrosine-phosphorylated proteins. In the past we used the anti-phosphotyrosine antibody, 4G10, from Millipore, a staple for these type of westerns. When presented with an opportunity to try a different antibody, APY03, from your company, we were eager to see the results. Overall, we were pleased with the consistent result when compared with the Millipore antibody. In addition, the most significant advantage between the two antibodies was the huge savings in pricing. With these two factors, reduction in cost and reliable results, we will continue to use your product and suggest it to others." Jeff McDermott and Gladis SanchezUniversity of Kansas Medical Center Customer Results Reviewed Product: Anti-Phosphotyrosine Mouse Monoclonal Antibody (Cat. # APY03) Application:Western blot “I have compared APY03 and 4G10 antibodies and I believe that APY03 is as good as 4G10 with the advantage of APY03 being that it is cheaper and also you get more western blots out of one purchase compared to 4G10.” Felipe Navarrete, University of Massachusetts Customer Results Western blot analysis of extracts from non-capacitated (N. Cap) and capacitated sperm (Cap). Cap extracts express a high level of proteins that are phosphorylated on tyrosine residues. Manufacturers’ protocols followed. Primary antibody dilution of APY03 was increased to 1:5000 and still produces a robust signal that detects phosphotyrosine residues undetected by 4G10. For the most recent publications citing this product, please contact our Technical Service department at tservice@cytoskeleton.com
Visit our Signal-Seeker™ Tech Tips and FAQs page for technical tips and frequently asked questions regarding this and other Signal-Seeker™ products click here If you have any questions concerning this product, please contact our Technical Service department at tservice@cytoskeleton.com
