Proteomic tools for quantitation by mass spectrometry

Authors


Abstract

  I.Introduction182
 II.Relative Quantitation184
 A.  2-D Gel Electrophoresis184
 B.  Metabolic Isotopic Labeling185
     1.  15N185
     2.  13C Enrichment and Depletion185
     3.  Select Isotopic Amino Acid Incorporation186
 C.  Chemical Labeling186
     1.  Labeling During Proteolysis: 18O Incorporation During Enzymatic Cleavage186
     2.  Isotopic Tags—Isotope-Codes Affinity Tag Reagents (ICAT)187
     3.  Isotopic Tags—Acid-Labile Isotope-Coded Extractants (ALICE)188
     4.  Lysine-Specific Labeling189
     5.  Phosphoserine- and Phosphothreonine-Specific Labeling189
     6.  N-Terminus Labeling189
         a.  Nicotinyl-N-hyxroxysuccinimide189
         b.  Acylation189
         c.  Amidination-quantitation using enhanced signal tags (QUEST)190
         d.  C-terminusu labeling190
 D.  Differential Mass Mapping190
III.Absolute Quantitation191
 IV.Choice of Instrumentation191
  V.Overview, Discussion, and Future Directions191
References 192

Techniques for the quantitation of proteins and peptides by mass spectrometry (MS) are reviewed. A range of labeling processes is discussed, including metabolic, enzymatic, and chemical labeling, and techniques that can be employed for comparative and absolute quantitation are presented. Advantages and drawbacks of the techniques are discussed, and suggestions for the appropriate uses of the methodologies are explained. Overall, the metabolic incorporation of isotopic labels provides the most accurate labeling strategy, and is most useful when an internal standard for comparative quantitation is needed. However, that technique is limited to research that uses cultured cells. © 2003 Wiley Periodicals, Inc., Mass Spec Rev 22:182–194, 2003; Published online in Wiley Interscience (www.interscience.wiley.com). DOI 10.1002/mas.10048

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