Automatic diagnosis of tuberculosis disease based on plasmonic ELISA and color-based image classification

Tuberculosis (TB) remains one of the most devastating infectious diseases and its treatment efficiency is majorly influenced by the stage at which infection with the TB bacterium is diagnosed. The available methods for TB diagnosis are either time consuming, costly or not efficient. This study employs a signal generation mechanism for biosensing, known as Plasmonic ELISA, and computational intelligence to facilitate automatic diagnosis of TB. Plasmonic ELISA enables the detection of a few molecules of analyte by the incorporation of smart nanomaterials for better sensitivity of the developed detection system. The computational system uses k-means clustering and thresholding for image segmentation. This paper presents the results of the classification performance of the Plasmonic ELISA imaging data by using various types of classifiers. The five-fold cross-validation results show high accuracy rate (>97%) in classifying TB images using the entire data set. Future work will focus on developing an intelligent mobile-enabled expert system to diagnose TB in real-time. The intelligent system will be clinically validated and tested in collaboration with healthcare providers in Malaysia

Development of sandwich-type electrochemical antibody and aptamer sensors for detection of Mycobacterium tuberculosis

Early diagnosis of Mycobacterium tuberculosis is very important to reduce the number of fatal cases and allow fast recovery. This can be done by using smear microscopy method; however, the interpretation of results requires skilled personnel due to tendency of the method to produce false-negative results (lack of sensitivity). Therefore, in this study, sandwich-type electrochemical immunosensor and aptasensor were compared for detection of CFP10-ESAT6 complex antigen as a biomarker of Mycobacterium tuberculosis, based on graphene/polyaniline (GP/PANI) modified screen-printed gold electrode (SPGE). GP/PANI nanocomposite was synthesized and characterized using Raman spectroscopy and Field Emission Scanning Electron Microscope (FESEM) in order to confirm the formation of nanocomposite. Then, it was dispersed in 3-aminopropyltriethoxysilane (APTES) and used as an electrode modifier. The morphology of the fabricated GP/PANI-SPGE was analyzed using FESEM and it showed a rough and porous surface while Energy Dispersive X-ray (EDX) spectroscopy has shown all the presence elements of GP/PANI on SPGE surface. Based on cyclic voltammetry (CV) characterization, the fabricated GP/PANI-SPGE has shown a large surface area compared to unmodified electrode. Capturing probes (anti­CFP10-ESAT6 antibodies or aptamer) were immobilized onto the surface of GP/PANI-SPGE. Iron/gold magnetic nanoparticles (Fe3Q4/Au MNPs) conjugated with anti-CFP10-ESAT6 was used to complete the sandwich system. The Fe3Q4/Au MNPs was synthesized and characterized using Ultraviolet-Visible (UV-Vis) spectrophotometer, High Resolution-Transmission Electron Microscopy (HR-TEM) and X-ray Diffraction (XRD) before conjugating it with anti-CFP10-ESAT6. Differential pulse voltammetry (DPV) technique was used to investigate the analytical performance of both immunosensor and aptasensor with its corresponding CFP10-ESAT6 antigen. The detection time was within 2 hours. Under optimum conditions, both sensors showed comparable results with almost the same limit of detection (LOO) of 1.47 ng/ml for immunosensor and 1.52 ng/ml for aptasensor. However, aptasensor showed better specificity and reproducibility than immunosensor. The methods developed from these processes were then integrated into a portable reader, which provides a good correlation with conventional methods on detection of M. tuberculosis in sputum samples. Henceforth, the developed biosensor demonstrated potential as a practical screening tool for M. tuberculosis detection

Enhancement of electrochemical properties using iron oxide-gold nanocomposite for tuberculosis detection based on rGO-APTES modified screen-printed electrode

A new electrochemical immunosensor based on iron oxide-gold nanocomposite (Fe 3 O 4 -Au NC) as signal enhancement was developed for sensitive detection of M. tuberculosis , the causative agent of tuberculosis (TB). Reduced graphene oxide-(3-aminopropyl)-triethoxysilane (rGO-APTES) modified screen-printed electrode (SPE) was employed as a matrix to immobilize the capture antibodies (Ab). The detection signal was amplified due to the large amounts of Ab captured on the large surface area of modified SPE. The Fe 3 O 4 -Au NC were functionalized as label to conjugate with primary antibodies (Ab 1 ). Due to the homogeneity and narrow size distribution of Fe 3 O 4 -Au NC, several Ab 1 were bound on each nanocomposite. The introduction of Fe 3 O 4 -Au-Ab 1 onto the electrode surface through sandwich immunoreactions enhances the performance of the sensor. The proposed immunosensor offered a sensitive detection of TB in the range of 0– 9.0 μg /mL, with a detection limit of 1.32 ng/mL. A disposable and sensitive SPE-based electrochemical immunoassay would serve as a promising platform to detect various mycobacterium pathogens for future diagnostic studies

Cellulose and vanadium plasmonic sensor to measure Ni2+ ions

A novel vanadium–cellulose composite thin film-based on angular interrogation surface plasmon resonance (SPR) sensor for ppb-level detection of Ni(II) ion was developed. Experimental results show that the sensor has a linear response to the Ni(II) ion concentrations in the range of 2–50 ppb with a determination coefficient (R2) of 0.9910. This SPR sensor can attain a maximum sensitivity (0.068° ppb−1), binding affinity constant (1.819 × 106 M−1), detection accuracy (0.3034 degree−1), and signal-to-noise-ratio (0.0276) for Ni(II) ion detection. The optical properties of thin-film targeting Ni(II) ions in different concentrations were obtained by fitting the SPR reflectance curves using the WinSpall program. All in all, the proposed Au/MPA/V–CNCs–CTA thin-film-based surface plasmon resonance sensor exhibits better sensing performance than the previous film-based sensor and demonstrates a wide and promising technology candidate for environmental monitoring applications in the future

Electrochemical immunosensor for detection of Mycobacterium tuberculosis

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

Aptasensor for the Detection of Mycobacterium tuberculosis in Sputum Utilising CFP10-ESAT6 Protein as a Selective Biomarker

A portable electrochemical aptamer-antibody based sandwich biosensor has been designed and successfully developed using an aptamer bioreceptor immobilized onto a screen-printed electrode surface for Mycobacterium tuberculosis (M. tuberculosis) detection in clinical sputum samples. In the sensing strategy, a CFP10-ESAT6 binding aptamer was immobilized onto a graphene/polyaniline (GP/PANI)-modified gold working electrode by covalent binding via glutaraldehyde linkage. Upon interaction with the CFP10-ESAT6 antigen target, the aptamer will capture the target where the nano-labelled Fe3O4/Au MNPs conjugated antibody is used to complete the sandwich format and enhance the signal produced from the aptamer–antigen interaction. Using this strategy, the detection of CFP10-ESAT6 antigen was conducted in the concentration range of 5 to 500 ng/mL. From the analysis, the detection limit was found to be 1.5 ng/mL, thereby demonstrating the efficiency of the aptamer as a bioreceptor. The specificity study was carried out using bovine serum albumin (BSA), MPT64, and human serum, and the result demonstrated good specificity that is 7% higher than the antibody–antigen interaction reported in a previous study. The fabricated aptasensor for M. tuberculosis analysis shows good reproducibility with an relative standard deviation (RSD) of 2.5%. Further analysis of M. tuberculosis in sputum samples have shown good correlation with the culture method with 100% specificity and sensitivity, thus making the aptasensor a promising candidate for M. tuberculosis detection considering its high specificity and sensitivity with clinical samples

Sandwich Electrochemical Immunosensor for Early Detection of Tuberculosis Based on Graphene/Polyaniline-Modified Screen-Printed Gold Electrode

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 tuberculosis biomarker 10-kDa culture filtrate protein (CFP10). The prepared GP/PANI nanocomposite was characterized using Fourier transform infrared spectroscopy (FTIR) and field emission scanning electron microscopy (FESEM). The chemical bonding and morphology of GP/PANI-modified SPGE were studied by Raman spectroscopy and FESEM coupled with energy dispersive X-ray spectroscopy, respectively. From both studies, it clearly showed that GP/PANI was successfully coated onto SPGE through drop cast technique. Cyclic voltammetry was used to study the electrochemical properties of the modified electrode. The effective surface area for GP/PANI-modified SPGE was enhanced about five times compared with bare SPGE. Differential pulse voltammetry was used to detect the CFP10 antigen. The GP/PANI-modified SPGE that was fortified with sandwich type immunosensor exhibited a wide linear range (20⁻100 ng/mL) with a low detection limit of 15 ng/mL. This proposed electrochemical immunosensor is sensitive, low sample volume, rapid and disposable, which is suitable for tuberculosis detection in real samples