Nanozymes in Point-of-Care Diagnosis: An Emerging Futuristic Approach for Biosensing

Abstract

Nanomaterial-based artificial enzymes (or nanozymes) have attracted great attention in the past few years owing to their capability not only to mimic functionality but also to overcome the inherent drawbacks of the natural enzymes. Numerous advantages of nanozymes such as diverse enzyme-mimicking activities, low cost, high stability, robustness, unique surface chemistry, and ease of surface tunability and biocompatibility have allowed their integration in a wide range of biosensing applications. Several metal, metal oxide, metal-organic framework-based nanozymes have been exploited for the development of biosensing systems, which present the potential for point-of-care analysis. To highlight recent progress in the field, in this review, more than 260 research articles are discussed systematically with suitable recent examples, elucidating the role of nanozymes to reinforce, miniaturize, and improve the performance of point-of-care diagnostics addressing the ASSURED (affordable, sensitive, specific, user-friendly, rapid and robust, equipment-free and deliverable to the end user) criteria formulated by World Health Organization. The review reveals that many biosensing strategies such as electrochemical, colorimetric, fluorescent, and immunological sensors required to achieve the ASSURED standards can be implemented by using enzyme-mimicking activities of nanomaterials as signal producing components. However, basic system functionality is still lacking. Since the enzyme-mimicking properties of the nanomaterials are dictated by their size, shape, composition, surface charge, surface chemistry as well as external parameters such as pH or temperature, these factors play a crucial role in the design and function of nanozyme-based point-of-care diagnostics. Therefore, it requires a deliberate exertion to integrate various parameters for truly ASSURED solutions to be realized. This review also discusses possible limitations and research gaps to provide readers a brief scenario of the emerging role of nanozymes in state-of-the-art POC diagnosis system development for futuristic biosensing applications.

Keywords: ASSURED diagnostics; Biosensing; Catalytic nanomaterials; Nanozymes; Point-of-care diagnosis.

Fig. 1

World Health Organization’s ASSURED criteria for point of care device

Fig. 2

A brief timeline of the development of nanozymes over the years (natural enzymes and artificial enzymes are listed for comparison)

Fig. 3

Schematic representation of different enzyme-mimicking activities by nanozymes in the presence of superoxide anions produced by single electron donor

Fig. 4

a Nanozyme-strip designed for the detection of Ebola virus. It shows standard colloidal gold strip, MNPs as nanozyme probe, which amplifies the signal by generating colour reaction and can be visualized by naked-eye. Specificity of the nanozyme probe towards EBOV-GP, but not for other virus proteins such as nucleoprotein of influenza A virus or New Bunya virus and the peroxidase-mimicking activity of antibody coated nanozymes towards different peroxidase substrates such as DAB, TMB, AEC [121]. Copyright 2015 Elsevier. b Paper-based bioassay for the detection of glucose using ceria NPs as nanozyme. Schematic representation of ceria nanoparticle-immobilized paper in APTS, colorimetric response of ceria-immobilized paper strips depending on the different range of H₂O₂ concentrations and linear calibration curve for H₂O₂ detection using ceria-immobilized paper strips [190]. Copyright 2011 American Chemical Society

Fig. 5

Different simple and effective conjugation chemistry for the surface modification of nanozymes with numerous biorecognition ligands

Fig. 6

a Schematic illustration of Pd–Pt nanozyme-based lateral flow assay for the detection of Escherichia coli O157:H7. TMB = 3,3′,5,5′-tetramethylbenzidine; NC = nitrocellulose; NP = nanoparticles [133]. Copyright 2018 Elsevier. c Schematic illustration of the strategy behind the Fe₃O₄ MNPs linked colorimetric aptasensors assay for the detection of thrombin [192]. Copyright 2010 Elsevier

Fig. 7

a Representation of the UA detection using PtNPs. It shows the preparation of cellulose strip consisting PtNPs and colour changes after addition of different concentration of UA [126]. Copyright 2019 Royal Society of Chemistry. b Principle behind the fluorescent-based detection of glucose using MIL-53(Fe) bifunctional nanozyme [200].Copyright 2018 Royal Society of Chemistry. c Schematic illustration of the principle of the CL sensor array based on the triple-channel properties of the luminol-functionalized Ag nanoparticles and H₂O₂ chemiluminescent (Lum AgNPs − H₂O₂ CL) system utilized for the detection of carbamate pesticides and organophosphorus [201]. Copyright 2015 American Chemical Society. d Schematic representation of the fabrication of an immunosensor based on Au@Pd@Au nanocluster nanozymes for the detection of PSA. Inset shows the differential pulse voltammetry (DPV) curve of different nanocomposites [204]. Copyright 2018 Elsevier. e Construction of an electrochemical immunosensor based on Au@Pt nanocluster nanozymes for the detection of PSA [203]. Copyright 2018 Elsevier

Fig. 8

Comparison of the catalytic efficiency for the peroxidase-like activity of different-shaped and different-sized AuNPs and natural enzyme (HRP) [212]. Copyright 2020 Javier Lou-Franco et al.

Fig. 9

a Schematic illustration for the preparation of nanohybrid system consisting Au–Ni (nickel)-based bimetallic nanoparticles doped in graphite carbon nitride sheets and its application for the selective and sensitive detection of glucose [218]. Copyright 2019 Elsevier B.V. b Schematic representation of Au@PtNP/GO microbeads preparation and Au@PtNP/GO nanozyme-based electrochemical POC device for quantitative detection of H₂O₂ [120]. Copyright 2019 Elsevier

Fig. 10

a AuNPs@MoS₂-QDs composite assisted catalytic oxidation of TMB in the presence of H₂O₂ and UV–Vis spectra of colorimetric sensing of glucose [219]. Copyright 2018 Elsevier. b Analysis of sensitivity and selectivity of glucose biosensing using Fe₃O₄@C yolk–shell nanostructured nanozymes. Schematic demonstration of simple and label-free colorimetric biosensing of glucose using Fe₃O₄@C nanocomposites. It shows the dose responsive curve, linear calibration curve and selectivity of the glucose detection assay [135]. Copyright 2017 Royal Society of Chemistry. c Enhanced catalytic rate/catalytic efficiency (peroxidase-like activity) of nanozymes dependence on different composition and shapes. d Schematic illustration and visual demonstration of inhibition of peroxidase-like activity of MoS₂ nanozyme in the presence of different concentrations of lipase. Inset demonstrate the enzyme-mimicking activity of MoS₂ with or without lipase [158]. Copyright 2018 Wiley–VCH

Fig. 11

a Schematic illustration of the colorimetric biosensing of Hg²⁺ ions using Au nanozyme PAD based on the mercury ion-assisted enhanced catalytic efficiency of Au nanozyme. In the presence of Hg²⁺ ions on the Au nanozyme PAD the catalytic oxidation of TMB enhanced significantly due to the formation of Au–Hg amalgam, resulting in generation of blue colour in the paper-chip. Photograph shows the Au nanozyme PAD with test samples consisting increasing concentration of Hg levels ranging from 0.2–2000 ng. Calibration curve shows the colorimetric response of Au nanozyme PAD-assisted TMB oxidation in the presence of Hg ions [222]. Copyright 2017 Han et al. b Colorimetric biosensor for the detection of proteolytic biomarkers based on unusual peroxidase-mimicking activity of Au nanozymes. Picture shows the naked-eye visualization of colorimetric response with the increasing concentration of protease (0–2.0 mg mL⁻¹) [217]. Copyright 2018 McVey et al.

Fig. 12

a Peroxidase-mimicking activity of H-Pt nanozyme-based ImmunoCAP (laboratory tests for serum allergen-specific IgE antibodies) diagnostic system for the sensitive detection of human IgE [132]. Copyright 2018 Royal Society of Chemistry. b Schematic representation of the fabrication of electrochemical immunosensor using magnetic silica NPs/GO nanocomposites for the detection of cancer antigen 153 (CA 153). Specificity assay of the electrochemical immunosensor for the detection of antigen 153 [227]. Copyright 2014 Elsevier

Fig. 13

a Schematic illustration of preparation of Pd-NPs/meso-C-based test strips and their application in quantitative detection of H₂O₂ using smartphone and APP [147]. Copyright 2018 Elsevier. b Schematic illustration of the preparation of the µPAD and assay procedure for the sensitive detection of carcinoembryonic antigen (CEA) [153]. Copyright 2018 Elsevier. c Schematic illustration of skin interstitial fluid (ISF) extraction using glucose-biosensing microneedle patch (GBMP) and the principle of colorimetric glucose detection using GOx-MnO2@GO as a colorimetric probe and smart phone-based app for quantitative analysis at hyperglycaemic condition [238]. Copyright 2020 Elsevier

Fig. 14

a Working mechanism of cobalt oxyhydroxide nanoflakes integrated paper-based point-of-care detection of acetylcholinesterase (AChE). The figure shows fabrication of the paper-based test strips and visual colorimetric detection of AChE based on the colour intensity on the test strips [124]. Copyright 2019 The Royal Society of Chemistry. b Schematic representation of the proposed biosensor using magnetic nanoparticles (MNps)-based immunoseparation, nanocluster signal amplification and quantitative detection of Salmonella typhimurium using smartphone-based application [239]. Copyright 2019 Elsevier

Fig. 15

a Overview of developed colorimetric and chronoamperometric assay for the detection of glucose using peroxidase-like activity of mesoporous iron oxide nanoparticles [242]. Copyright 2017 American Chemical Society. b Schematic representation of the assay for the detection of tumour-associated plasma (and serum) p53 autoantibody. A neutravidin-modified screen-printed carbon electrode was functionalized with biotinylated p53. Serum/plasma samples containing p53-specific autoantibody were then incubated with IgG/Au–NPFe₂O₃NC nanocatalysts. Au–NPFe₂O₃NC catalysed the oxidation of TMB in the presence of H₂O₂ and produced a blue-coloured complex product (naked eye). The level of p53 autoantibody was detected via measuring the intensity (UV–Vis) and amperometric current generated by the product [134]. Copyright 2017 American Chemical Society. c Schematic representation of a paper electrode-based flexible pressure sensor for point-of-care immunoassay with digital multimeter readout. Photographs of flexible pressure sensor, bendability of flexible pressure sensor, and immune device based on flexible pressure sensor [165]. Copyright 2018 American Chemical Society

Fig. 16

a Comparison of two different glucose detection model through cascade reaction. Conventional approach for colorimetric glucose detection using enzyme: glucose oxidase (GO) and horseradish peroxidase (HRP). Glucose detection using a synthetic bifunctional nanozyme (graphitic carbon nitride- g-C3N4: GCN). Photocatalytic aerobic oxidation of glucose leads to the production of H₂O₂ further oxidized by bifunctional nanozyme to produce colorimetric signal [246]. Copyright 2019 Nature. b Simplified representation of the assay for detection of Pseudomonas aeruginosa (PA) using Au nanozymes modified with F-23 aptamer. There is suppression of peroxidase-like activity of Au nanozymes in the presence of F-23 aptamer; however, the activity resumed in the presence of PA. The electrochemical detection of TMB reduction on screen-printed carbon electrode in the presence of PA is shown [247]. Copyright 2019 Springer Nature