Electrochemical immunosensor for simultaneous determination of interleukin-1 beta and tumor necrosis factor alpha in serum and saliva using dual screen printed electrodes modified with functionalized double–walled carbon nanotubes

Abstract

Dual screen-printed carbon electrodes modified with 4-carboxyphenyl-functionalized double-walled carbon nanotubes (HOOC-Phe-DWCNTs/SPCEs) have been used as scaffolds for the preparation of electrochemical immunosensors for the simultaneous determination of the cytokines Interleukin-1β (IL-1β) and factor necrosis tumor α (TNF-α). IL-1β. Capture antibodies were immobilized onto HOOC-Phe-DWCNTs/SPCEs in an oriented form making using the commercial polymeric coating Mix&Go™. Sandwich type immunoassays with amperometric signal amplification through the use of poly-HRP-streptavidin conjugates and HO as HRP substrate and hydroquinone as redox mediator were implemented. Upon optimization of the experimental variables affecting the immunosensor performance, the dual immunosensor allows ranges of linearity extending between 0.5 and 100 pg/mL and from 1 to 200 pg/mL for IL-1β and TNF-α, respectively, these ranges being adequate for the determination of the cytokines in clinical samples. The achieved limits of detection were 0.38 pg/mL (IL-1β) and 0.85 pg/mL (TNF-α). In addition, the dual immunosensor exhibits excellent reproducibility of the measurements, storage stability of the anti-IL-Phe-DWCNTs/SPCE and anti-TNF-Phe-DWCNTs/SPCE conjugates, and selectivity as well as negligible cross-talking. The dual immunosensor was applied to the simultaneous determination of IL-1β and TNF-α in human serum spiked at clinically relevant concentration levels and in real saliva samples.

An impedimetric molecularly-imprinted biosensor for Interleukin-1β determination, prepared by in-situ electropolymerization on carbon screen-printed electrodes

“…Among these, carbon supports (SPCEs) are very popular, being highly effective and of low cost [8]. As far as we know, there is a single electrochemical biosensor work published for IL-1β, which is an immunosensor assembled on double walled carbon nanotubes showing limits of detection of 0.38 pg/mL [9]. Yet, this biosensor uses commercial antibodies as biorecognition element, which increases the cost of the overall process, and decreases the stability of the device and its lifetime.…”

Printed Electrodes in Microfluidic Arrays for Cancer Biomarker Protein Detection

“…The same strategy was later used for transforming growth factor-β (TGF-β) detection with LOD of 10 pg mL −1 [ 110 ]. They further modified the array with 4-carboxyphenyl-functionalized double-walled carbon nanotubes to improve the detection for multiplex proteins, TNF-α and IL-1β, with LODs 0.85 and 0.38 pg mL −1 , respectively [ 111 ]. Meanwhile, our group recently published a multiplexed detection assay of four prostate cancer biomarkers, PSA, ETS-related gene protein (ERG), insulin-like growth factor-1 (IGF-1), and VEGF-D, using a system with strep-poly-HRP80 coupled with SPCE from Kanichi Ltd. modified with the glutathione-AuNP layers from Mani et al [ 93 ].…”

Multiplexed Electrochemical Immunosensors for Clinical Biomarkers

“…Calibration plots with ranges of linearity extending between 0.5 and 100 pg·mL −1 and from 1 to 200 pg·mL −1 for IL-1β and TNF-α, respectively, were obtained with limits of detection of 0.38 pg·mL −1 (IL-1β) and 0.85 pg·mL −1 (TNF-α). The dual immunosensor was validated by application to the simultaneous determination of both cytokines in human serum spiked at clinically relevant concentration levels and in real saliva samples [28]. Related with cytokines determination, Baraket et al [29] prepared an electrochemical immunosensor using silicon chip technology for real time detection of various interleukins: IL-1β, IL-10, and IL-6 which were secreted in acute stages of inflammation.…”