Bimodal determination of immunoglobulin E by fluorometry and ICP-MS by using platinum nanoclusters as a label in an immunoassay

Abstract

The authors describe the use of platinum nanoclusters (PtNCs) as bimodal labels in a competitive immunoassay for immunoglobulin E (IgE). Both fluorometry and inductively coupled plasmamass spectrometry (ICP-MS) are used. Optimization of the PtNCs synthesis process using lipoic acid as ligand was carried out. The time for synthesis and the effect of NaOH added to the PtNCs precursor mixture was optimized with the aim to obtain PtNCs with strong fluorescence and low size dispersity. Maximal fluorescence was obtained at excitation/emission wavelengths of 455/620 nm. The average diameter (1.5 nm) and crystal structure (face-centered cubic structure) of the PtNCs were determined by HR-TEM. It was calculated that each PtNC contains 116 Pt atoms at average. Labelling of the antibody (Ab) against IgE with PtNCs was optimized in terms of recognition capabilities and fluorescence signal. A molar ratio (Ab:PtNCs) of 1:11 is found to be best. A competitive immunoassay for IgE was developed and detection was carried out by using both ICP-MS (by measuring 195 Pt) and fluorometry. The limit of detection (LOD) of the fluoroimmunoassay is 0.6 ng mL-1 of IgE. The LOD of the ICP-MS method is as low as 0.08 ng mL-1. The method was evaluated by analyzing four (spiked) serum samples by ICP-MS. No sample pretreatment excepting dilution is needed. Results compared favorably with those obtained by a commercial ELISA kit.

Multiplex bioimaging of proteins-related to neurodegenerative diseases in eye sections by laser ablation - Inductively coupled plasma – Mass spectrometry using metal nanoclusters as labels

“…Next, MNCs were characterised by fluorescence, HR-TEM and ICP-MS measurements. HR-TEM images showed in the three cases MNCs with a spherical metal nucleus and FCC crystal structure [26,29,30]. As described elsewhere [26,29,30], the average particle size determined for each type of MNCs was: 1.5 ± 0.6 nm for PtNCs, 2.20 ± 0.04 nm for AuNCs, and 2.9 ± 0.1 nm for AgNCs.…”

“…The syntheses of the three thiolated MNCs (PtNCs, AuNCs and AgNCs) were carried out following the protocols described elsewhere [26,29,30]. A similar bottom-up procedure was used in all cases: the metal precursor (salt of the metal with a high oxidation state) is dissolved in basic aqueous media (adjusted with a NaOH solution) and subsequently reduced by NaBH4 under the presence of the stabilising agent (lipoic acid).…”

“…The labelling of the specific primary Ab (anti-h-MT1/2, anti-h-CFH or anti-h-APP) was first performed with PtNCs as the elemental label to accomplish the individual bioimaging distribution of the proteins in three consecutive eye tissue sections from the same donor. The procedure was carried out following the carbodiimide chemistry, and it was previously optimised in terms of keeping the properties of both components (i.e., the recognition capabilities of the Ab and the fluorescence properties of the PtNCs) [29]. 198 L of PtNCs solution was mixed with 100 μL of each primary Ab (10 μg mL −1 ), keeping a 1:20 molar ratio Ab:PtNCs.…”

“…In a previous publication describing the determination of IgE in human sera using PtNCs by conventional nebulisation ICP-MS [29], the molar ratio between the Ab and PtNCs was optimised. Such optimisation was performed to maximise the amount of metal per Ab to achieve high amplification for ICP-MS detection while keeping the recognition capabilities of the Ab.…”

Synthesis of Size Monodisperse Water-Soluble Metal Nanoclusters for Protein Quantification by Elemental Mass Spectrometry

“…MNCs (AuNCs, AgNCs and PtNCs) have been recently reported as elemental tags for the determination of proteins in serum samples and biological tissues by using ICP-MS (i.e., serum) and laser ablation (LA) coupled to ICP-MS (i.e., tissues and cultured cells). For example, PtNCs were used as bimodal labels in a competitive immunoassay for immunoglobulin E determination in serum samples [5]. The huge amplification achieved by ICP-MS detection gave rise to a better limit of detection than the fluoroimmunoassay and the commercial ELISA kit.…”