Ubiquitin

Applications:\nSuitable for use in ELISA, Western Blot, Immunohistochemistry and Immunoflourescence. Other applications not tested.\n\nRecommended Dilution:\nELISA: 0.1-1ug/ml\nWestern Blot: 1-3ug/ml; Prepare lysis buffer in 10mM N-ethylmaleimide to inhibit ubiquitin-conjugating enzymes. N-ethylmaleimide inactivates certain enzymes by blocking free sulfhydryls. After electrophoresis and transfer, pre-incubate transferred membranes in denaturing buffer (6M guanidine-HCl 20mM Tris-HCl, pH 7.5, 5mM b-mercaptoethanol, 1mM PMSF) for 30-60 minutes at 4 degrees C, followed by extensive PBS washing.\nImmunohistochemistry: Alcohol/paraformaldhyde-fixed paraffin-embedded or Frozen tissue sections. Requires heat induced epitope retrieval with citrate buffer (pH, 6.0) for formalin-fixed, paraffin-embedded sections. To stain, incubate for 30-60 minutes at RT or overnight at 4 degrees C.\nImmunofluorescence: 2-5ug/ml\nOptimal dilutions to be determined by the researcher.\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20 degrees C. Aliquots are stable for 12 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

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SPECIFICATIONS

Catalog Number

U1000-04B

Size

100ug

Applications

ELISA, IF, IHC, WB

Hosts

Mouse

Reactivities

Hum, Mouse, Bov, Ch/Bird, Dro/Arth

Form

Supplied as a liquid in PBS, pH 7.4, 0.1% sodium azide.

P Type

Mab

Purity

Purified by Protein A affinity chromatography.

Isotype

IgG1,k

References

1. Manetto, V., et al., PNAS 85: 4501-4505 (1988). 2. Shaw, G. & Chau, V., PNAS 85: 2854-2858 (1988). 3. Askanas, V., et al., Neurosci. Lett. 130: 73-76 (1991). 4. Schmidt, M.L., et al., Am. J. Pathol. 139: 53-65 (1991). 5. Mayer, R.J., et al., Ann. NY Acad. Sci. 674: 149 (1992). 6. Hirano, S., et al., Cell 70: 293-301 (1992). 7. Sternsdorf, T., et al., J. Cell Biol. 139(7): 1621-1634 (1997). 8. Kim, T-W., et al., J. Biol. Chem. 272: 11,006-11,010 (1997). 9. Cuervo, A.M., et al., Mol. Biol. Cell 9: 1995-2010 (1998). 10. Verdier, F., et al., J. Biol. Chem. 273(43): 28,185-28,190 (1998). 11. Laroia, G., et al., Science 284(5413): 499-502 (1999). 12. Marti, A., et al., Nature Cell Biol. 1(1): 14-19 (1999). 13. Sternsdorf, T., et al., Mol. Cell Biol. 19(7): 5170-5178 (1999). 14. Mullally, J.E., et al., J. Biol. Chem. 276(32):3 0,366-30,373 (2001). 15. Ma, Q. & Baldwin, K.T., J. Biol. Chem. 275(12): 8432-8438 (2000). 16. Fernandez-Funez, P., et al., Nature 408: 101-106 (2000). 17. Holbert, S., et al., PNAS 98(4): 1811-1816 (2001). 18. Ruffner, H., et al., PNAS 98(9): 5134-5139 (2001). 19. Roig, J., et al., PNAS 97(26): 14,346-14,351 (2000).

Additional Info

Recognizes bovine ubiquitin, both conjugated and unconjugated. Reacts with a single chain 8.5kD protein. The ubiquitin molecule appears to be present in all eukaryotic cells and has an identical primary structure in all animals. Ubiquitin is present in the nucleus, cytoplasm, and on cell surface membranes. Species Crossreactivity: human, chicken, mouse, C. elegans and Drosophila.