Phos-tag beads as an immunoblotting enhancer for selective detection of phosphoproteins in cell lysates
Analytical Biochemistry Volume 389 Issue 1
Page 83-85
published_at 2009-06-01
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| File | |
| Title ( eng ) |
Phos-tag beads as an immunoblotting enhancer for selective detection of phosphoproteins in cell lysates
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| Creator |
Kinoshita-Kikuta Emiko
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| Source Title |
Analytical Biochemistry
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| Volume | 389 |
| Issue | 1 |
| Start Page | 83 |
| End Page | 85 |
| Abstract |
The low specificity of anti-phosphoprotein antibodies is often a problem in immunoblotting analyses. We introduce a simple pretreatment procedure for cell lysates to give more-specific detection of phosphoproteins in immunoblotting. Cellular phosphoproteins were preferentially trapped on Phos-tag agarose phosphate-affinity beads in a homemade spin-centrifuge microtube unit, and nonphosphorylated proteins were excluded in the filtrate. The phosphoprotein-bound beads suspended in a sample-loading dye solution were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by Western blotting. We demonstrated improved detection of phosphorylated Shc and mitogen-activated protein kinase isoforms in A431 cell lysates by this new technique
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| Keywords |
Phos-tag
Immunoblotting enhancer
Anti-phosphoprotein antibody
Phosphoprotein
Phosphorylation
Signal transduction
Western blotting
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| NDC |
Biology [ 460 ]
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| Language |
eng
|
| Resource Type | journal article |
| Publisher |
Elsevier
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| Date of Issued | 2009-06-01 |
| Rights |
Copyright (c) 2009 Elsevier Inc.
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| Publish Type | Author’s Original |
| Access Rights | open access |
| Source Identifier |
[ISSN] 0003-2697
[ISSN] 1096-0309
[DOI] 10.1016/j.ab.2009.03.026
[NCID] AA00524867
[NCID] AA11537838
[DOI] http://dx.doi.org/10.1016/j.ab.2009.03.026
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