Dual Colorimetric/Electrochemical Detection of Salmonella typhimurium Using a Laser-Induced Graphene Integrated Lateral Flow Immunoassay Strip.

Foodborne illnesses caused by the ingestion of contaminated foods or beverages are a serious concern due to the millions of reported cases per year. It is essential to develop sensitive and rapid detection methods of foodborne pathogens to ensure food safety for producers and consumers. Unfortunately, current detection techniques still suffer from time-consuming operations and the need for highly skilled personnel. Here, we introduce a highly sensitive dual colorimetric/electrochemical detection approach for Salmonella enterica serovar typhimurium (S. typhimurium) based on a laser-induced graphene-integrated lateral flow immunoassay (LIG-LFIA) strip. The LIG electrode was fabricated by laser engraving on a polyimide tape containing a pseudo silver/silver chloride reference electrode from silver sintering and chlorination. Using double-sided tape inserted into the strip, automatic sequential reagent delivery was enabled for the dual-mode signal readout by single-sample loading. A gold-deposited gold nanoparticle strategy was first employed to simultaneously obtain a colorimetric signal for early screening and a signal turn-on electrochemical response for high-sensitivity and -quantitative analysis. A superior performance of the strip was established, characterized by a short analysis time (12 min assay +15 min sample preparation), a broad working concentration range (1 cfu/10 mL to 108 cfu/mL), and the lowest limit of detection (1 ± 0.5 cfu/10 mL; mean ± standard deviation, n = 3) among reported multimode S. typhimurium detection schemes. The strip was successfully applied in the analysis of various food products without any bacterial enrichment or amplification required, and the results were comparable to those of the standard culture method.

Automated Paper-Based Femtogram Sensing Device for Competitive Enzyme-Linked Immunosorbent Assay of Aflatoxin B1 Using Submicroliter Samples.

Microfluidic paper-based analytical devices (µPADs) are promising biosensors that may be used in a variety of bioanalytical applications. A µPAD for automating the competitive enzyme-linked immunosorbent assay (ELISA) of small-sized target detection at the femtogram level using submicroliter samples is reported in this study. The proposed µPAD was integrated with a sucrose valve to automate the sequential delivery of reagents, providing simple control of reagent delivery time and simple operation. The use of a sample solution dropping location at the zones on the device that had been prepared with an antibody-conjugated enzyme before immersion in a running buffer allowed minimization of sample volume to 0.6 µL, while eliminating the possible loss of a target molecule by adsorption on the membrane, thus improving detection sensitivity. Furthermore, the proposed device was successfully applied to the automation of competitive ELISA for the detection of aflatoxin B1 (AFB1), a potent carcinogen that causes substantial health risks to humans worldwide, with a detection limit of 60 femtograms or 0.1 ng/mL. The method developed in this study provides high sensitivity, small sample volume, on-site and equipment-free measurements, low-cost operation, and user-friendliness. This approach could be used to analyze small-sized molecules in the fields of food safety and quality control, environmental monitoring, and clinical diagnostics.

A paper-based conductive immunosensor for the determination of Salmonella Typhimurium.

We report for the first time a highly sensitive and rapid quantitative method for the detection of Salmonella Typhimurium (S. Typhimurium) using a conductive immunosensor on a paper-based device (PAD). S. Typhimurium monoclonal antibodies (MA) were first immobilized on a paper-based device and then captured by S. Typhimurium. After an immunoreaction on the device, the polyclonal antibody-colloidal gold conjugate (PA-AuNPs) was dropped to bind with S. Typhimurium. After a complete sandwich reaction, a dark red color appeared on the paper-based device, which can be observed by the naked eye for a rapid screening test. The electrical conductivity of PA-AuNPs between the screen-printed electrodes on the paper-based device was also measured for an accurate quantitative analysis. The electrical conductivity correlated well with the concentration of S. Typhimurium, which was controlled by the amount of S. Typhimurium attached to the polyclonal antibody-colloidal gold conjugate. The device showed a linear correlation for the concentration of the S. Typhimurium in the range of 10-108 CFU mL-1 in a logarithmic plot, with an R2 value of 0.9882 and a limit of detection (LOD) as low as 10 CFU mL-1. This simple, highly sensitive, and rapid method for the S. Typhimurium detection was successfully performed within 30 min, and it can be developed into small portable measuring devices in order to facilitate preliminary screening tests.

A nanocomposite prepared from platinum particles, polyaniline and a Ti3C2 MXene for amperometric sensing of hydrogen peroxide and lactate.

A nanocomposite consisting of platinum particles, polyaniline and Ti3C2 MXene (Pt/PANI/MXene) was used to modify a screen-printed carbon electrode (SPCE) to obtain sensors for hydrogen peroxide and lactate. This nanocomposite was characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM) and X-ray powder diffraction (XRD) to determine the physical morphologies and the nanocomposite elements. The modified electrode exhibited the improved current response towards hydrogen peroxide (H2O2) compared with an unmodified electrode and provided a low detection limit of 1.0 µM. When lactate oxidase was immobilized on the modified electrode, the electrode responded to lactate via the H2O2 generated in the enzymatic reaction. The lactate assay was performed by amperometry at a constant potential of +0.3 V (vs. Ag/AgCl). The linear range was found to be from 0.005 to 5.0 mM with a detection limit of 5.0 µM for lactate. Ultimately, this biosensor was used for the determination of lactate in milk samples with high stability and reliability. Graphical abstractSchematic representation of a novel platinum particles/polyaniline/MXene nanocomposite (Pt/PANI/MXene) for screen-printed carbon electrode (SPCE) modification to enhance the specific surface area for immobilization of lactate oxidase (LOx) and use as enzymatic biosensor for lactate determination in milk sample.

Long-term stabilization of hydrogen peroxide by poly(vinyl alcohol) on paper-based analytical devices.

Stabilizing reagents that can be deposited onto paper is an important issue for researchers who depend on paper-based analytical devices (PADs), because long-term stability of the devices is essential in point-of-care testing. Here, we found that poly(vinyl alcohol) (PVA) would stabilize hydrogen peroxide placed on a paper substrate following exposure to air. Horseradish peroxidase was employed as a sample in colorimetric measurements of PADs after hydrogen peroxide and 3,3',5,5'-tetramethylbenzidine were deposited as substrates in an enzymatic reaction. The addition of PVA to hydrogen peroxide significantly suppressed its degradation. Concentrations of PVA that ranged from 0.5 to 2%, increased the duration of the stability of hydrogen peroxide, and the results for a PVA concentration of 1% approximated those of 2% PVA. Storage of the PADs at 4 °C in a refrigerator extended the stability of the hydrogen peroxide containing 2% PVA by as much as 30 days. The stability of hydrogen peroxide without PVA was degraded after one day under room temperature.

Analysis of sudan I, sudan II, sudan III, and sudan IV in food by HPLC with electrochemical detection: Comparison of glassy carbon electrode with carbon nanotube-ionic liquid gel modified electrode.

A method was developed for analyzing of sudan I, sudan II, sudan III, and sudan IV by high performance liquid chromatography coupled with an electrochemical detector. The electrochemical oxidation of each compound was investigated with a carbon nanotube-ionic liquid gel modified glassy carbon (MWNTs-IL-Gel/GC) electrode using cyclic voltammetry. The results were compared with those of glassy carbon electrodes. At the MWNTs-IL-Gel/GC electrode, highly reproducible, well-defined cyclic voltammograms for sudan oxidation were obtained. In the flow system, MWNTs-IL-Gel/GC exhibited highly reproducible and well-defined amperometric signals without peak tailing. In addition, the determination of sudan dyes by mean of isocratic reverse-phase HPLC, with amperometric detection at MWNTs-IL-Gel/GC and GC electrode, have been investigated. The chromatograms showed excellent separation. The detection limits ranged from 0.001 to 0.005ppm. The method was sensitive, selective and could be applicable for the assay of sudan dyes in soft drink samples.

Polyaniline/graphene quantum dot-modified screen-printed carbon electrode for the rapid determination of Cr(VI) using stopped-flow analysis coupled with voltammetric technique.

Polyaniline/graphene quantum dots (PANI/GQDs) were used to modify a screen-printed carbon electrode (SPCE) in a flow-based system. A method for rapidly determining the Cr(VI) concentrations by using stopped-flow analysis has been developed using an Auto-Pret system coupled with linear-sweep voltammetry using the PANI/GQD-modified SPCE. The GQDs, synthesized in a botton-up manner from citric acid, were mixed with aniline monomer in an optimized ratio. The mixture was injected into an electrochemical flow cell in which electro-polymerization of the aniline monomer occurred. Under conditions optimized for determining Cr(VI), wide linearity was obtained in the range of 0.1-10 mg L(-1), with a detection limit of 0.097 mg L(-1). For a sample volume of 0.5 m L, the modified SPCE can be used continuously with a sample-throughput of more than 90 samples per hour. In addition, this proposed method was successfully applied to mineral water samples with acceptable accuracy, and the quantitative agreement was accomplished in deteriorated Cr-plating solutions with a standard traditional method for Cr(VI) detection.

Electrochemical detection of human papillomavirus DNA type 16 using a pyrrolidinyl peptide nucleic acid probe immobilized on screen-printed carbon electrodes.

An electrochemical biosensor based on an immobilized anthraquinone-labeled pyrrolidinyl peptide nucleic acid (acpcPNA) probe was successfully developed for the selective detection of human papillomavirus (HPV) type 16 DNA. A 14-mer acpcPNA capture probe was designed to recognize a specific 14 nucleotide region of HPV type 16 L1 gene. The redox-active label anthraquinone (AQ) was covalently attached to the N-terminus of the acpcPNA probe through an amide bond. The probe was immobilized onto a chitosan-modified disposable screen-printed carbon electrode via a C-terminal lysine residue using glutaraldehyde as a cross-linking agent. Hybridization with the target DNA was studied by measuring the electrochemical signal response of the AQ label using square-wave voltammetric analysis. The calibration curve exhibited a linear range between 0.02 and 12.0 µM with a limit of detection and limit of quantitation of 4 and 14 nM, respectively. This DNA sensing platform was successfully applied to detect the HPV type 16 DNA from a PCR amplified (240 bp fragment of the L1 gene) sample derived from the HPV type 16 positive human cancer cell line (SiHa), and failed to detect the HPV-negative c33a cell line. The sensor probe exhibited very high selectivity for the complementary 14 base oligonucleotide over the non-complementary oligonucleotides with sequences derived from HPV types 18, 31 and 33. The proposed sensor provides an inexpensive tool for the early stage detection of HPV type 16, which is an important biomarker for cervical cancer.

Highly sensitive determination of cadmium and lead using a low-cost electrochemical flow-through cell based on a carbon paste electrode.

A low-cost thin-layer electrochemical flow-through cell based on a carbon paste electrode (CPE), was constructed for the highly sensitive determination of cadmium(II) (Cd(2+)) and lead(II) (Pb(2+)) ions. The sensitivity of the proposed cell for Cd(2+) and Pb(2+) ion detection was improved by using the smallest channel height without the need for any complicated electrode modification. Under the optimum conditions, the detection limits of Cd(2+) and Pb(2+) ions (0.08 and 0.07 µg dm(-3), respectively) were 13.8- and 11.4-fold lower than that of a commercial flow cell (1.1 and 0.8 µg dm(-3), respectively). Moreover, the percentage recoveries of Cd(2+) and Pb(2+) for the in-house designed thin-layer flow cell were higher than those for the commercially available cell in all tested water samples, and within the acceptable range. The proposed flow cell is promising as an inexpensive and alternative one for the highly sensitive monitoring of heavy metal ions.

Graphene-carbon paste electrode for cadmium and lead ion monitoring in a flow-based system.

An environment friendly electrode for determining Cd(2+) and Pb(2+) levels in an automated flow system was successfully developed. Cyclic voltammetry and square wave anodic stripping voltammetry (SWASV) coupled with sequential injection analysis (SIA) were employed to study the electrochemical behavior of the electrode. The in situ bismuth-modified graphene-carbon paste electrode (Bi-GCPE) exhibited excellent electrooxidation of Cd(2+) and Pb(2+) in the automated flow system with a significantly higher peak current for both metal ions compared with the unmodified CPE. The limits of detection from this method were 0.07 and 0.04 µg L(-1) for Cd(2+) and Pb(2+), respectively, with a linear oxidation peak current response for Cd(2+) and Pb(2+) in the range of 0.10-50.0 µg L(-1) under optimum conditions. The Bi-GCPE was also applied for the determination of Cd(2+) and Pb(2+) in low- (tap water) and high- (sea bass fish and undulated surf clam tissues) matrix complexity samples by automated flow system. The recoveries were acceptable and ranged from 70.4% to 120% for Cd(2+) and 65.8% to 113.5% for Pb(2+).