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http://hdl.handle.net/2445/123187
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: https://doi.org/10.1002/elps.201500495 |
It is part of: | Electrophoresis, 2016, vol. 37, p. 1220-1231 |
URI: | http://hdl.handle.net/2445/123187 |
Related resource: | https://doi.org/10.1002/elps.201500495 |
ISSN: | 0173-0835 |
Appears in Collections: | Articles publicats en revistes (Enginyeria Química i Química Analítica) |
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File | Description | Size | Format | |
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655669.pdf | 1.04 MB | Adobe PDF | View/Open |
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