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Curbing tuberculosis (TB) requires a combination of good strategies, including a proper prevention measure, diagnosis, and treatment. This study proposes an improvised tuberculosis diagnosis based on an amperometry approach for the sensitive detection of MPT64 antigen in clinical samples. An MPT64 aptamer specific to the target antigen was covalently attached to the carboxyphenyl diazonium-functionalized carbon electrode via carbodiimide chemistry. The electrochemical detection assay was adapted from a sandwich assay format to trap the antigen between the immobilized aptamer and horseradish peroxidase (HRP) tagged polyclonal anti-MPT64 antibody. The amperometric current was measured from the catalytic reaction response between HRP, hydrogen peroxide, and hydroquinone, which is used as an electron mediator. From the analysis, the detection limit in the measurement buffer was 1.11 ng mL-1. Additionally, the developed aptasensor exhibited a linear relationship between the current signal and the MPT64 antigen-spiked serum concentration ranging from 10 to 150 ng mL-1 with a 1.38 ng mL-1 detection limit. Finally, an evaluation using the clinical sputum samples from both TB (+) and TB (-) individuals revealed a sensitivity and specificity of 88% and 100%, respectively. Based on the analysis, the developed aptasensor was found to be simple in its fabrication, sensitive, and allowed for the efficient detection and diagnosis of TB in sputum samples.

Early diagnosis is highly crucial for life-saving and transmission management of tuberculosis (TB). Despite the low sensitivity and time-consuming issues, TB antigen detection still relies on conventional smear microscopy and culture techniques. To address this limitation, we report the development of the first amperometric dual aptasensor for the simultaneous detection of Mycobacterium tuberculosis secreted antigens CFP10 and MPT64 for better diagnosis and control of TB. The developed sensor was based on the aptamers–antibodies sandwich assay and detected by chronoamperometry through the electrocatalytic reaction between peroxidase-conjugated antibodies, H2O2, and hydroquinone. The CFP10 and MPT64 aptamers were immobilized via carbodiimide covalent chemistry over the disposable dual screen-printed carbon electrodes modified with a 4-carboxyphenyl diazonium salt. Under optimized conditions, the aptasensor achieved a detection limit of 1.68 ng mL−1 and 1.82 ng mL−1 for CFP10 and MPT64 antigens, respectively. The developed assay requires a small sample amount (5 µL) and can be easily performed within 2.5 h. Finally, the dual aptasensor was successfully applied to clinical sputum samples with the obtained diagnostic sensitivity (n = 24) and specificity (n = 13) of 100%, respectively, suggesting the readiness of the developed assay to be used for TB clinical application.

Aptamers are short single-stranded oligonucleotides (either DNA or RNA) that can fold into well-defined three-dimensional (3D) spatial structures which enable them to capture their specific target by complementary shape interactions. Aptamers are selected from large random libraries through the SELEX process and only a small fraction of the sequence is involved in direct docking with the target. In this paper, we describe the possible truncation variants of zearalenone (ZEA) aptamer which might be an effective binding region for the target. The originally selected zearalenone (ZEA) aptamer was 80-mer in length and shown to bind the target with a high affinity (Kd = 41 ± 5 nM). Herein, computational docking simulation was performed with 15 truncated variants to determine the predicted binding energy and responsible binding site of the aptamer-analyte complex. The results revealed that 5 truncated variants had binding energy lower than - 7.0 kcal/mol. Circular dichroism analysis was performed on the shortlisted aptamer and the conformational change of aptamers was observed with the presence of an analyte. Aptamer Z3IN (29-mer) was chosen as the most enhanced affinity for its target with a dissociation constant of 11.77 ± 1.44 nM. The aptamer was further applied in the electrochemical aptasensor of ZEA based on an indirect competitive format. The results demonstrated that the truncated aptamer leads to an enhancement of the sensitivity of the biosensor.

An electrochemical aptasensor is described for determination of the phytohormone of zearalenone (ZEA). The gold electrode was modified with ZEA via covalent attachment using cysteamine-hydrochloride and 1,4-phenylene diisocyanate linker. A truncated ZEA aptamer with a dissociation constant of 13.4 ± 2.1 nM was used in an aptasensor. The electrochemical property was investigated using square wave voltammetry for monitoring the change in the electron transfer using the ferro/ferricyanide system as redox probe. Under optimal experimental conditions, the response was best measured at a potential of 0.20 V (vs. Ag/AgCl). The signals depended on the competitive mechanism between the immobilised ZEA and free ZEA for the aptamer binding site. The aptasensor works in the range 0.01 to 1000 ng·mL−1 ZEA concentration, with a detection limit of 0.017 ng· mL−1. High degree of cross-reactivity with the other analogues of ZEA was observed, whereas none towards other mycotoxins. The aptasensor was further applied for the determination of ZEA in the extract of maize grain and showed good recovery percentages between 87 and 110%.

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis. Detection and control of infectious diseases is a major problem, especially in developing countries. Lateral flow immunoassay (LFIA) has been introduced as a handheld immunoassay-based point-of-care platform for an automated detection of TB. The CFP10-ESAT6 antigen of M. tuberculosis was used as the target in early detection of TB using LFIA strip-based POC strategy. An interesting platform based on optical signals is implemented as a colour change in the detection area that is visible to the naked eye. The gold nanoparticles (AuNPs) were used as the colour probe for the detection of a target of interest. The high-resolution transmission electron microscopy (HRTEM) image and ultraviolet-visible spectrophotometer (UV-Vis) analysis confirmed that the synthesized AuNPs were appropriate for the immunoassay designed. The platform consists of AuNPs conjugated with specific antibodies (Ab) to capture the antigen of M. tuberculosis. Under the capillary effect, sandwich immunoreactions of AuNP-Ab-antigen were performed on the test pad of the immunostrip, which can be observed by the colour signal on the test line of the strip with a short assay time. Furthermore, the newly developed biosensor was utilized in CFP10-ESAT6 antigen detection in human sputum specimens with satisfactory results. The characteristic coloured bands enable visual detection (naked eye) of target analyte without instrumentation. This noninvasive diagnose system which is sputum-based detection could provide user-friendly and affordable diagnostic tests in developing countries.

This work presents a simple green synthesis of gold nanoparticles (AuNPs) by using an aqueous extract of Etlingera elatior (torch ginger). The metabolites present in E. elatior, including sugars, proteins, polyphenols, and flavonoids, were known to play important roles in reducing metal ions and supporting the subsequent stability of nanoparticles. The present work aimed to investigate the ability of the E. elatior extract to synthesise AuNPs via the reduction of gold (III) chloride hydrate and characterise the properties of the nanoparticles produced. The antioxidant properties of the E. elatior extract were evaluated by analysing the total phenolic and total flavonoid contents. To ascertain the formation of AuNPs, the synthesised particles were characterised using the ultraviolet-visible (UV-Vis) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, high-resolution transmission electron microscopy (HRTEM), energy-dispersive X-ray (EDX) microscopy, and dynamic light scattering (DLS) measurement. The properties of the green synthesised AuNPs were shown to be comparable to the AuNPs produced using a conventional reducing agent, sodium citrate. The UV-Vis measured the surface plasmon resonance of the AuNPs, and a band centered at 529 nm was obtained. The FTIR results proved that the extract contained the O-H functional group that is responsible for capping the nanoparticles. The HRTEM images showed that the green synthesized AuNPs were of various shapes and the average of the nanoparticles’ hydrodynamic diameter was 31.5 ± 0.5 nm. Meanwhile, the zeta potential of −32.0 ± 0.4 mV indicates the high stability and negative charge of the AuNPs. We further successfully demonstrated that using the green synthesised AuNPs as the nanocomposite to modify the working surface of screen-printed carbon electrode (SPCE/Cs/AuNPs) enhanced the rate of electron transfer and provided a sensitive platform for the detection of Cu(II) ions.

Lateral flow immunoassays (LFIAs) are advantageous over conventional detection methods in terms of their simplicity and rapidity. These assays have been reported using various types of labels but colloidal gold nanoparticles are still the preferred choice as a label because of their easy synthesis, visual detection and stability. Tuberculosis, or TB, is an infectious bacterial disease caused by Mycobacterium tuberculosis. İt remains one of the deadliest diseases in the world. The detection of Mycobacterium tuberculosis using LFIAs was developed and analyzed using gold nanoparticle.

A rapid and sensitive sandwich electrochemical immunosensor was developed based on the fabrication of the graphene/polyaniline (GP/PANI) nanocomposite onto screen-printed gold electrode (SPGE) for detection of Mycobacterium tuberculosis (M. tuberculosis) antigen. The chemical bonding and morphology of GP/PANI-modified SPGE were studied by Raman spectroscopy and field enhance scanning electron microscopy-energy dispersive X-ray spectroscopy (FESEM-EDX), respectively. From both studies, it clearly showed that GP/PANI was successfully coated onto SPGE through drop cast technique. Cyclic voltammetry (CV) was used to study the electrochemical properties of the modified electrode. The effective surface area for GP/PANI-modified SPGE was enhanced when compared with bare SPGE. Differential pulse voltammetry (DPV) was used to detect the M. tuberculosis antigen. This proposed electrochemical immunosensor is sensitive, low sample volume, rapid, and disposable, which is suitable for tuberculosis detection in real samples.