title: Quantitation of surface-bound proteins on biochips using MALDI-TOF MS.
abstract: We report on a novel method for the quantitation of proteins specifically bound on a ligand-presenting biochip by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS).  The bound protein was digested by trypsin, and the resulting peptide fragments were analyzed by MALDI-TOF MS in the presence of an isotope-labeled internal standard (IS).  The IS has the same sequence as a reference peptide (RP) of the target protein digest, but a different molecular weight.  The absolute amount of the specifically bound protein on a biochip is then quantitated by comparison of mass intensities between the RP and the IS.  Because they have the same molecular milieu, the mass intensities of these two analytes represent the real amounts of analytes on the chip.  As a model system, we tested glutathione s-transferase (GST) and a GST-fusion protein, which were captured on glutathione-presenting biochips.  We observed that the glutathione densities on biochips showed a good correlation with the absolute quantity of the proteins.  We believe that our method will provide an alternative to currently existing tools for the absolute quantitation of surface-bound proteins.
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, Seoul, Korea.
authors.forename: Juhee
authors.initials: J
authors.lastname: Lee
authors.forename: Soo-Ryoon
authors.initials: SR
authors.lastname: Ryoo
authors.forename: Sang Kyung
authors.initials: SK
authors.lastname: Kim
authors.forename: Joong-Hoon
authors.initials: JH
authors.lastname: Ahn
authors.forename: Dal-Hee
authors.initials: DH
authors.lastname: Min
authors.forename: Woon-Seok
authors.initials: WS
authors.lastname: Yeo
mesh_terms: D002247:Carbon Isotopes / Q000032:analysis / Q000737:chemistry; D002264:Carboxylic Acids / Q000737:chemistry; D005978:Glutathione / Q000737:chemistry; D005982:Glutathione Transferase / Q000032:analysis*; D006046:Gold / Q000737:chemistry; D007553:Isotope Labeling; D008301:Maleimides / Q000737:chemistry; D008565:Membrane Proteins / Q000032:analysis*; D008970:Molecular Weight; D009587:Nitrogen Isotopes / Q000032:analysis / Q000737:chemistry; D040081:Protein Array Analysis / Q000295:instrumentation*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation* / Q000379:methods / Q000592:standards
chemical_list: D002247:Carbon Isotopes; D002264:Carboxylic Acids; D008301:Maleimides; D008565:Membrane Proteins; D009587:Nitrogen Isotopes; D006046:Gold; D005982:Glutathione Transferase; D005978:Glutathione