Description
General description
Puromycin is an aminonucleoside antibiotic, derived from the Streptomyces alboniger bacterium, that functions as a protein synthesis inhibitor that blocks translation through premature chain termination in the ribosome. Monoclonal antibodies to puromycin may be used with standard immunochemical methods to directly monitor translation, a method known as surface sensing of translation (SUnSET). Part of the molecule resembles the 3′ end of the aminoacylated tRNA, making it useful for protein translation analysis. Puromycin induces DNA fragmentation in thymocytes and in human HL-60 leukemia cells.
Puromycin is incorporated in neosynthesized proteins. In the presense of Puromycin only, this antibody detects Puromycin-incorporated neosynthesized proteins at multiple molecular weights. However, a decrease in signal is observed in the co-presense of Cycloheximide, an inhibitor of protein biosynthesis in eukaryotic organisms.
Application
Anti-Puromycin antibody, clone 4G11 detects puromycin incorporated into protein. Monoclonal antibodies to puromycin may also be used with standard immunochemical methods.
Immunocytochemistry Analysis: A 1:2,000 dilution from a representative lot detected Puromycin-incorporated neosynthesized proteins in HeLa cells treated with Puromycin. However, a decrease in signal is observed with the addition of Cycloheximide, an inhibitor of protein biosynthesis in eukaryotic organisms.
Western Blotting Analysis: A representative lot from an independent laboratory detected Puromycin-incorporated neosynthesized proteins in WB (David, A., et al. (2012). J Cell Biol. 197(1):45-57.; Schmidt, E., K., et al. (2009). Nat Methods. 6(4):275-277.).
Immunofluorescence Analysis: A representative lot from an independent laboratory detected Puromycin-incorporated neosynthesized proteins in IF (David, A., et al. (2012). J Cell Biol. 197(1):45-57.; Schmidt, E., K., et al. (2009). Nat Methods. 6(4):275-277.).
Fluorescence Activated Cell Sorting Analysis: A representative lot from an independent laboratory detected Puromycin-incorporated neosynthesized proteins in FACS (David, A., et al. (2012). J Cell Biol. 197(1):45-57.; Schmidt, E., K., et al. (2009). Nat Methods. 6(4):275-277.).
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Biochem/physiol Actions
Demonstrated to react with Human test sample, preincubated with Puromycin. Predicted to react with all species when test sample is incubated with Puromycin.
Physical form
Format: Purified
Analysis Note
Evaluated by Western Blotting in HEK293 cell lysates treated with Puromycin and Cyclohexamide, or with Puromycin only.
Western Blotting Analysis: A 1:12,500 dilution of this antibody detected Puromycin-incorporated neosynthesized proteins in HEK293 cell lysates treated with Puromycin only. This antibody also detected small mounts of puromycin in HEK293 cells treated with Puromycin and Cyclohexamide.
Legal Information
ALEXA FLUOR is a trademark of Life Technologies
