Eric Hamlett

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Publications (2)0 Total impact

  • Article: Analysis of protein posttranslational modifications using DIGE-based proteomics.
    Robert M DeKroon, Jennifer B Robinette, Cristina Osorio, Joseph S Y Jeong, Eric Hamlett, Mihaela Mocanu, Oscar Alzate
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    ABSTRACT: Difference gel electrophoresis (DIGE) is most often used to assess relative changes in the expression levels of individual proteins in multiple complex samples, and this information is valuable in making inferences about relative protein activity. However, a protein's activity is not solely dependent upon its expression level. A change in activity may also be influenced by myriad posttranslational modifications (PTMs), including palmitoylation, ubiquitination, oxidation, and phosphorylation. In this chapter, we describe the use of DIGE to determine specific PTMs by introducing specific labels or changes in pI and/or molecular weight.
    Methods in molecular biology (Clifton, N.J.) 01/2012; 854:129-43.
  • Article: Analysis of proteins using DIGE and MALDI mass spectrometry.
    Witold M Winnik, Robert M Dekroon, Joseph S Y Jeong, Mihaela Mocanu, Jennifer B Robinette, Cristina Osorio, Nedyalka N Dicheva, Eric Hamlett, Oscar Alzate
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    ABSTRACT: Difference gel electrophoresis (DIGE) is a common technique for characterizing differential protein expression in quantitative proteomics. Usually a combination of enzymatic digestion and peptide analysis by mass spectrometry is used to identify differentially expressed proteins following separation and statistical analysis by DIGE. In this chapter, methods for gel spot picking, enzymatic digestion, and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) for protein identification of DIGE-analyzed proteins are discussed. Two examples are given: first, a specific protein is used to test the sensitivity of the 2D DIGE/MALDI MS combination for protein quantification and identification, and second, several proteins with and without the labels typically used in DIGE are identified to demonstrate that these labels do not alter MS-based protein identification. Technical variations of protein gel spot preparation, in-gel digestion, and mass spectral protein identification are discussed.
    Methods in molecular biology (Clifton, N.J.) 01/2012; 854:47-66.
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