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High-throughput protein characterization using mass spectrometric immunoassay.

Urban A Kiernan·Kemmons A Tubbs·Karl Gruber·Dobrin Nedelkov·Eric E Niederkofler·Peter Williams·Randall W Nelson
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Abstract

A high-throughput mass spectrometric immunoassay system for the analysis of proteins directly from plasma is reported. A 96-well format robotic workstation was used to prepare antibody-derivatized affinity pipette tips for subsequent use in the extraction of specific proteins from plasma and deposition onto 96-well format matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) targets. Samples from multiple individuals were screened with regard to the plasma protein transthyretin (TTR), followed by analysis of the same plasma samples for the transthyretin-associated transport protein, retinol-binding protein (RBP). Analyses were able to detect the presence of posttranslationally modified TTR and RBP, as well as a mutation present in the TTR of one individual. Subsequent analyses of wild-type and mutated TTR using enzymatically active MALDI-TOF MS targets were able to identify the site and nature of the point mutation. The approach represents a rapid (approximately 100 samples/2 h, reagent preparation-to-data) and accurate means of characterizing specific proteins present in large numbers of individuals for proteomic and clinical/diagnostic purposes.

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Antibody AffinityBlood ProteinsHumansImmunoassayMalePrealbuminRetinol-Binding ProteinsRetinol-Binding Proteins, PlasmaSpectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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Urban A Kiernan, Kemmons A Tubbs, Karl Gruber, Dobrin Nedelkov, Eric E Niederkofler, Peter Williams, Randall W Nelson. 2002-02-01. High-throughput protein characterization using mass spectrometric immunoassay.. https://doi.org/10.1006/abio.2001.5478

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