Electrochemiluminescence (ECL) immunosensor for detection of Francisella tularensis on screen-printed gold electrode array

Abstract

An electrochemiluminescence (ECL) immunosensor for the rapid detection of the Francisella tularensis pathogen using whole antibodies or antibody fragments as capture biomolecule is described. A sandwich immunoassay was used with either lipopolysaccharide (LPS) or the whole inactivated bacterial cell (LVS) as a target, while Ru(bpy)3 (2+)-encapsulated silicate nanoparticles were linked to the secondary antibody and used as ECL labels. The assay was performed in a fluidic chip housed in a custom-built black box incorporating electronics, optics and fluidics. The obtained limit of detection for LPS was 0.4 ng/mL, while for the LVS it was 70 and 45 bacteria/mL when the capturing molecule was the whole antibody and the antibody F(ab) fragment, respectively.

Electrochemiluminescence Biosensors Using Screen-Printed Electrodes

“…Among nanomaterials employed to support [Ru(bpy) 3 ] 2+ as ECL label, silica nanoparticles are very common. The immunosensor for whole cells of Francisella tularensis detection (responsible for tularemia infectious disease) is an illustrative example [ 69 ]. In this work, silica-encapsulated [Ru(bpy) 3 ] 2+ nanoparticles are synthesized and coated with polydiallyldimethylammonium.…”

Electrochemical Immuno- and Aptamer-Based Assays for Bacteria: Pros and Cons over Traditional Detection Schemes

“…Most convenient are sandwich immunoassays with a secondary Ab labelled with Ru(II) tris(bipyridine) complex emitting light after electrochemical stimulation with such co-reactant as tripropylamine [ 163 , 164 ] (analogues of e-ELISA in which enzyme labels are replaced by a chemiluminescent reagent). 45 CFU mL −1 of Francisella tularensis with Ab fragments as capture biomolecules could be detected in a fluidic chip with screen-printed 42 Au electrodes array within a 30 min procedure [ 164 ], and down to 2.3 CFU mL −1 of E. coli O157:H7 were detected in 2 h with the automated electroluminescent 48-well singleplex plate sensor (250 µL samples) [ 163 ]. Additional 4 h sample pre-enrichment by ultrafiltration of 10 mL samples further decreased the LOD to 0.12 CFU mL −1 [ 163 ].…”

“…The existing commercial ECL-enabling analyzers, such as Roche cobas ® 6000 analyzers, allow from 170 to 2170 test per h, but they are not yet adapted for bacterial sensing [ 169 ] and may be not suitable for direct analysis in the blood. More portable devices are nevertheless being developed [ 164 ], though their sensitivity should be further improved.…”

Recent Advances in Electrochemiluminescence Sensors for Pathogenic Bacteria Detection

“…Antibodies are large Y-shaped proteins and are often applied as recognition elements to capture bacteria. A well-known example is ELISA (enzyme-linked immunosorbent assay), in which the specific reaction between the target pathogen and the antibody connects the pathogen to the enzyme (horse radish peroxidase, HRP) and then produces a color reaction through the enzyme and the substrate [58,68,69,70]. When HRP is replaced with Ru complex and the capture antibody modified on a working electrode, the limits of detection (LODs) observed for ELISA and the ECL immunoassay for BoNT/A were 12 and 3 pg/mL, and for BoNT/B, they were 17 and 13 pg/mL, respectively.…”