Phosphothreonine

Reversible protein phosphorylation plays a central role in numerous biochemical pathways and functions to alter protein activity and/or conformation (3,7). Methods for detecting protein phosphorylation have predominately relied upon the use of radioactive 32P for either the in vitro or in vivo phosphorylation reaction (2,6). To identify the specific amino acid residues that become phosphorylated on a specific protein, phosphoamino acid analysis is then required (2,3,6,7). The use of radioactivity and the multi-step analysis process which follows makes the entire process both hazardous and tedious. Over the past decade, antibodies specific for phosphotyrosine were developed, many of which can detect a single phosphorylated tyrosine residue (1,4,5). Although antibodies to phosphotyrosine have been relatively straight forward to produce, the extensive structural similarity between phosphoserine and phosphothreonine has contributed to the difficulty in raising highly specific antibodies to these phosphoamino acids (6).\n\nApplications: \nSuitable for use in ELISA, Western Blot, and Immunoprecipitation . Other applications not tested.\n\nRecommended Dilution:\nELISA: 0.5ug/ml (kinase assay) \nWestern Blot: 2ug/ml versus mouse brain lysate. Incubate at RT for 4 hours.\nOptimal dilutions to be determined by the researcher.\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. For long-term storage, aliquot and store at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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SPECIFICATIONS

Catalog Number

P4077-10

Size

100ug

Applications

ELISA, IP, WB

Hosts

Mouse

Form

Supplied as a liquid in PBS, pH 7.4, 0.1% sodium azide, 40% glycerol.

P Type

Pab

Purity

Purified by immunoaffinity chromatography.

Isotype

IgG2a,k

References

1. Frackelton, A.R., et al., Mol. Cell. Biol. 3: 1343 (1983). 2. Edelman, A.M., et al., Annu. Rev. Biochem. 56: 567 (1987). 3. Hunter, T., Cell 50: 823 (1987). 4. Glenny, J.R., et al., J. Immunological Meth. 109: 277 (1988). 5. Sengupta, A., et al., Proc. Natl. Acad. Sci. USA 85: 8062 (1988). 6. Heffetz, D., et al., Methods in Enzymology 210: 44 (1991). 7. Hunter, T., Methods in Enzymology 200: 3 (1991). 8. Rintam

Additional Info

Recognizes Phosphothreonine, peptidyl-phosphothreonine, and threonine-phosphorylated proteins. Binding of Ly-pT-Gly-HRP (0.2ug/100ul/well) to an anti-phosphotherine coated microplate (0.5ug/100ul/well) is inhibited by 95% with the addition of 2-10ug/ml anti-phosphothreonine. No inhibition is evident by inhibition of phosphoserine, phosphotyrosine or theonine at concentrations of up to 1mg/ml. Species crossreactivity: species independent.