Current Advances in Paper-Based Biosensor Technologies for Rapid COVID-19 Diagnosis

Abstract

The global coronavirus disease 2019 (COVID-19) pandemic has had significant economic and social impacts on billions of people worldwide since severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first reported in Wuhan, China, in November 2019. Although polymerase chain reaction (PCR)-based technology serves as a robust test to detect SARS-CoV-2 in patients with COVID-19, there is a high demand for cost-effective, rapid, comfortable, and accurate point-of-care diagnostic tests in medical facilities. This review introduces the SARS-CoV-2 viral structure and diagnostic biomarkers derived from viral components. A comprehensive introduction of a paper-based diagnostic platform, including detection mechanisms for various target biomarkers and a COVID-19 commercial kit is presented. Intrinsic limitations related to the poor performance of currently developed paper-based devices and unresolved issues are discussed. Furthermore, we provide insight into novel paper-based diagnostic platforms integrated with advanced technologies such as nanotechnology, aptamers, surface-enhanced Raman spectroscopy (SERS), and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas. Finally, we discuss the prospects for the development of highly sensitive, accurate, cost-effective, and easy-to-use point-of-care COVID-19 diagnostic methods.

Keywords: COVID-19; Lateral flow assay (LFA); Paper-based biosensors; Point-of-care testing (POCT); SARS-CoV-2.

Fig. 1

SARS-CoV-2 genome, structure comprising major proteins, and variation levels of biomarkers across the duration of the infection. a The SARS-CoV-2 genome codes ten genes, and the genes are arranged in the sequence 5′ cap structure-ORF1/ab-spike (S)-envelope (E)-membrane (M)-nucleocapsid (N)-3′ poly (A) tail. Primer–probe sets for SARS-CoV-2 RNA amplification developed by research groups around the world [US Centers for Disease Control and Prevention (US-CDC) – target sequences: 28,287 ~ 28,358, 28,681 ~ 28,752, 29,164 ~ 29,230; China CDC – target sequences: 13,342 ~ 13,460, 28,881 ~ 28,979; Charité–Universitätsmedizin Berlin in Germany (Charité) – target sequences: 15,431 ~ 15,530, 26,269 ~ 26,381; National Institute of Infectious Disease in Japan (Japan NIID) – target sequence: 29,125 ~ 29,282; and University of Hong Kong (HKU) – target sequences: 18,778 ~ 18,909, 29,145 ~ 29,254]. b SARS-CoV-2 is mainly composed of four major proteins: spike (S) (red), membrane (M) (orange), envelope (E) (green), and nucleocapsid (N) (purple) proteins. c Temporal dynamics of the viral load and antigen and antibody levels. Since the types and amounts of biomarkers present in a patient's body fluid differ depending on the stage of infection, it is critical to select an appropriate biomarker and a method that can effectively detect it for an accurate diagnosis of COVID-19

Fig. 2

Paper-based diagnostic platforms including device components, detection mechanisms for different target markers, a antigens, b antibodies, and c RNA. a To detect the SARS-CoV-2 antigens, a specific antibody pair is required. These capture and detection antibodies detect SARS-CoV-2-specific antigens (S and N proteins) while forming a sandwich complex. After 15–20 min of sample loading, the appearance of color in the test and control lines is confirmed visually or by a portable analyzer. b In serological tests (detecting IgM and IgG antibodies), the N (or S) proteins of SARS-CoV-2 are conjugated with gold nanoparticles and used as signal molecules to detect IgM and IgG antibodies. Anti-human IgM (or IgG) antibodies are immobilized on a nitrocellulose membrane to form test lines. When the sample contains the SARS-CoV-2-specific IgM or IgG antibodies, the antibodies are bound to the N (or S) protein-conjugated gold nanoparticles and finally bound to the test line, resulting in vivid color. c Isothermal amplification techniques combined with an LFA contribute to achieving POC tests for SARS-CoV-2 RNA detection. First, an isothermal amplification process is performed for target RNA amplification, and then an LFA reaction is performed so that the results can be easily checked

Fig. 3

a Cellular receptor (ACE2)-based LFA for detecting SARS-CoV-2 S1 antigen, reproduced with permission from [63], copyright 2021 Elsevier. b Development of scFv-Fc-based LFA for detection of the SARS-CoV-2 N protein. Highly sensitive and specific scFv-Fc fusion proteins are rapidly screened by phage display technology, reproduced with permission from [62], copyright 2021 Elsevier. c Configuration of detecting system to quantify LFA results with the photon-counting approach and representative results for IgG antibody detection (concentrations range: from 1000 to 0.1 ng/mL), reproduced with permission from [74], copyright 2020 AIP. d LFA strip to detect anti-SARS-CoV-2 IgA antibody and the simple and universal smartphone reader to detect the optical signal from LFA, reproduced with permission from [75], copyright 2021 Elsevier. e Lateral Flow Strip Membranes (LFSM)-based on highly specific and sensitive detection of SARS-CoV-2. The LFSM assay allows simultaneous detection of the multiple regions of SARS-CoV-2 RNA in a sing test, reproduced with permission from [77, 76], copyright 2020 ACS. f Principle of reverse transcription-enzymatic recombinase amplification (RT-ERA). The RT-ERA has the capability of ultrasensitive, field-deployable, and simultaneous dual-gene detection of SARS-CoV-2 RNA, reproduced with permission from [79], copyright 2020 Springer nature

Fig. 4

a PFST-μPADs for quantitative SARS-CoV-2 IgA/IgM/IgG assay, reproduced with permission from [81], copyright 2021 ACS. b Three-dimensional μPADs for detecting SARS-CoV-2 specific antibodies based on affinity between cellulose and cellulose binding domain, reproduced with permission from [87], copyright 2021 ACS. c A label-free ePAD for detecting SARS-CoV-2-specific IgG and IgM antibodies, reproduced with permission from [56], copyright 2021 Elsevier. d A new ePAD-based COVID-19 diagnosis using ZnO NW-enhanced working electrode, reproduced with permission from [90], copyright 2021 Elsevier

Fig. 5

Development trends of EUA-approved commercialized antigen (a) and serological tests (b) for the diagnosis of COVID-19. a To satisfy the demand for high-throughput testing, LFA-type diagnostic tests have been mainly developed. For rapid, convenient, and cost-effective diagnosis, the kits are visually readable and developed for testing using nasal swab samples. b LFA-based serology kits accounted for only 28% of total products due to the low sensitivity. Most LFA-based serology tests visually confirm the results and target two or more antibodies

Fig. 6

a Fabrication process of the dual-mode SiO2@Au@QD and schematic of dual-mode LFA for detecting anti-SARS-CoV-2 IgM and IgG antibodies, reproduced with permission from [99], copyright 2020 ACS. b Lanthanide-doped nanoparticles-based LFA for detecting anti-SARS-CoV-2 IgG, reproduced with permission from [103], copyright 2020 ACS. c DNA aptamers-based LFA for detecting SARS-CoV-2 N antigen, reproduced with permission from [111], copyright 2020 RSC. d SERS-based LFA for SARS-CoV-2-specific IgM detection, reproduced with permission from [122], copyright 2021 ACS. e Workflow of CRISPR-based DETECTR assay. The SARS-CoV-2 DETECTR comprises RNA extraction, RT-LAMP, Cas12 detection, and LFA, reproduced with permission from [129], copyright 2020 Nature research