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Title: Analysis of serum transthyretin by on-line immunoaffinity solid-phase extraction capillary electrophoresis mass spectrometry using magnetic beads
Author: Peró Gascón, Roger
Pont Villanueva, Laura
Benavente Moreno, Fernando J. (Julián)
Barbosa Torralbo, José
Sanz Nebot, María Victoria
Keywords: Electroforesi capil·lar
Espectrometria de masses
Camps magnètics
Capillary electrophoresis
Mass spectrometry
Magnetic fields
Issue Date: 2016
Publisher: Wiley-VCH
Abstract: In this paper, an on-line immunoaffinity solid-phase extraction capillary electrophoresis mass spectrometry (IA-SPE-CE-MS) method using magnetic beads (MBs) is described for the analysis of serum transthyretin (TTR), which is a protein related to different types of amyloidosis. First, purification of TTR from serum was investigated by off-line immunoprecipitation and CE-MS. The suitability of three Protein A (ProA) MBs (Protein A Ultrarapid AgaroseTM (UAPA), Dynabeads® Protein A (DyPA) and SiMAG-Protein A (SiPA)) and AffiAmino Ultrarapid AgaroseTM (UAAF) MBs to prepare an IA sorbent with a polyclonal antibody (Ab) against TTR, was studied. In all cases results were repeatable and it was possible the identification and the quantitation of the relative abundance of the 6 most abundant TTR proteoforms. Although recoveries were the best with UAPA MBs, UAAF MBs were preferred for on-line immunopurification because Ab was not eluted from the MBs. Under the optimised conditions with standards in IA-SPE-CE-MS, microcartridge lifetime (>20 analyses/day) and repeatability (2.9 and 4.3 % RSD for migration times and peak areas) were good, the method was linear between 5- 25 µg·mL-1 and limit of detection (LOD) was around 1 µg·mL-1 (25 times lower than by CE-MS, 25 µg·mL-1). A simple off-line sample pretreatment based on precipitation of the most abundant proteins with 5% (v/v) of phenol was necessary to clean-up serum samples. The potential of the on-line method to screen for familial amyloidotic polyneuropathy type I (FAP-I), which is the most common hereditary systemic amyloidosis, was demonstrated analysing serum samples from healthy controls and FAP-I patients.
Note: Versió postprint del document publicat a:
It is part of: Electrophoresis, 2016, vol. 37, p. 1220-1231
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ISSN: 0173-0835
Appears in Collections:Articles publicats en revistes (Enginyeria Química i Química Analítica)

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