Translating in vitro diagnostics from centralized laboratories to point-of-care locations using commercially-available handheld meters

Abstract

There is a growing demand for high-performance point-of-care (POC) diagnostic technologies where in vitro diagnostics (IVD) is fundamental for prevention, identification, and treatment of many diseases. Over the past decade, a shift of IVDs from the centralized laboratories to POC settings is emerging. In this review, we summarize recent progress in translating IVDs from centralized labs to POC settings using commercially available handheld meters. After introducing typical workflows for IVDs and highlight innovative technologies in this area, we discuss advantages of using commercially available handheld meters for translating IVDs from centralized labs to POC settings. We then provide comprehensive coverage of different signal transduction strategies to repurpose the commercially-available handheld meters, including personal glucose meter, pH meter, thermometer and pressure meter, for detecting a wide range of targets by integrating biochemical assays with the meters for POC testing. Finally, we identify remaining challenges and offer future outlook in this area.

Graphical Abstract

1. Introduction: Current trends in in vitro diagnostics: from centralized labs to POC setting

The general public has a growing demand for high-performance point-of-care (POC) diagnostic technologies where in vitro diagnostics (IVD) is fundamental for prevention, identification, and treatment of chronical, infectious and terminal diseases, in addition to maintaining healthy lives [1]. For example, qualitative or quantitative detections of clinically relevant biomarkers, such as small molecules, proteins, and nucleic acids, is of great importance to improve the diagnosis as well as to provide timely and effective treatment for heart disease [2], bacterial infection [3] or cancers in advanced stages [4]. Among the detection methods, biosensors, which is typically consists of a biorecognition element to bind its target and a signal transduction element to convert the binding event into a physiochemical detectable signal, play a dominant role, because of high affinity and selectivity of biorecognition elements, such as antibody-antigen, DNA/RNA hybridization, enzyme-substrate, and host-guest interactions for a wide range of biomarkers and other targets [5]. In addition, recent advances in biomedical research, nanotechnology, and engineering, have contributed to various signal transduction methods, including the use of electrochemical [6], fluorescent [7], colorimetric [8] or magnetic signal [9], as well as those methods using glucose [10], temperature [11], pH [12], time [13], distance [14], or pressure [15] as readout. These emerging techniques have continuously propelled the shift of IVDs from the traditional centralized laboratories that generally require expensive instruments, sophisticated sampling steps, and highly skills technicians, to POCTs where portable devices can be used by users with minimal or no training to carry out IVDs in resource limited settings, including remote areas and home [16, 17]. These POC tests (POCTs) are becoming increasingly important to meet the demand of personalized medicine [18].

The IVDs around the world are still dominated by analyses carried out in centralized labs, in which many automated analytical processes are emerging to enable the simultaneously analysis of various samples. While these central clinical labs can provide sensitive and selective diagnostic assays, such as high-throughput immunoassays, quantitative polymerase chain reaction (q-PCR (or RT-PCR)), flow cytometry, and mass spectrometry tests, they are often time-consuming, labor intensive and require sophisticated instruments and trained operators. In some cases, these limitations usually result in long turnaround time that not only delay the diagnosis but also prevent the effective monitoring of postoperative recovery. To overcome these limitations, many POCTs have been developed to improve patient care in many aspects of IVDs, such as capability of rapid detection of infection, personalized disease treatment, and self-management of chronic diseases and therapies beyond centralized lab and in many decentralized locations [19], such as the doctor’s office, or a remote clinical lab. While many of these cheaper and smaller POC tests provide a faster turnaround, many challenges remain before their delivery to end patient users for home testing. From the patient’s perspective, the increased desire for self-health management drives the need for the development of user-friendly, and reliable POC devices for telemedicine.

In this review we summarize recent progress in translating IVDs from centralized labs to POC settings using commercially available handheld meters. We first briefly introduce the typical workflow for IVDs and highlight several new innovative technologies in this topic. Next, we discuss advantages of using commercially available handheld meters for translating IVDs from centralized labs to POC settings. We then provide a comprehensive coverage of different signal transduction strategies to repurpose the commercially-available handheld meters, including personal glucose meter (PGM), pH meter, thermometer and pressure meter, for the detection of a wide range of targets by integrating biochemical assays with these commercially-available meters for POC testing. Finally, we offer our assessment of the future outlook and remaining challenges in POCTs.

2. General workflow for IVDs

From operation point of view, the IVDs of biological samples generally consists of four basic steps (Figure 1): (a) sample preparation including extraction, separation, and purification from raw biological samples such as blood, urine, or saliva; (b) target recognition via different interactions with tailored specificity; (c) transduction of recognition events into a readable qualitative or quantitative signal; (d) signal amplification of the binding events to increase detection sensitivity. To allow the extensive use of IVDs in medical applications, considerable efforts have been made to explore innovative solutions to improve the outcomes of these steps.

Figure 1.

Schematic illustration of typical steps and strategies of in vitro diagnostics of biological samples.

2.1. Sample preparation

In IVDs, the sample preparation is an initial step to obtain analytical targets suitable for later steps in IVDs [20]. This step is extremely important for IVD of some biomarkers with low abundance in complex matrix. Based on the diversity of samples and targets, a variety of strategies for sample preparation have been developed, and extensively reviewed in the past few years [20–23]. Taking nucleic acid targets in blood samples as an example, traditional laboratory methods include physiochemical lysis, centrifugation, and solid phase extraction and isolation [20]. Despite progress, few of these methods are available for IVDs in the POC setting, mainly due to the multiple centrifuge and wash steps. To address this issue, microfluidics-based technology has been introduced that can integrate multiple sampling processes into one miniaturized unit [24, 25]. Some lab-on-a-chip (LOC) systems or lab-in-a-drop (LID) systems have also been developed [24], and demonstrated great potentials in IVDs. For example, Bandodkar et al. [26] developed a battery-free, wireless electronic sensing platform that integrates chronometric microfluidic chips with embedded colorimetric sensors, which could simultaneously monitor multiple targets, such as sweat rate/loss, lactate, pH, chloride, and glucose. Microfluidic paper-based analytical devices (μPADs) have emerged in recent years as a promising candidate to address the growing need for integrated sampling process [27]. Using this approach, Govindarajan et al. [28] reported a microfluidic origami paper for POC extraction of bacterial DNA from raw viscous samples. The core idea of this μPADs-based sample preparation device is sequencing complex chemical and physical processing steps via sequential folding of a flat substrate pieces. This method offers the possibility of POC, room temperature, DNA extraction, followed by storage of the extraction device at room temperature for delivery to more centralized diagnostic facilities. In addition to these approaches, readers interested in the general topic of sample preparation can refer to recent relevant reviews [20–23].

2.2. Target recognition

Once a biological sample has been prepared for an IVD test, it is analyzed by various techniques, including direct chemical or physical characterization and assays depending on affinity agents. Direct chemical or physical characterization involves instrumentations, including chromatographic equipment (e.g., HPLC and GC) and spectrometers. Various IVD assays will require affinity agents that specifically recognize the target; and today, target recognition in IVD assays most commonly includes antibody, enzymes, nucleic acid molecules and synthetic organic molecules. Among IVD assays, antibody-based immunoassays are the most common due to the wide range of targets being recognized with high selectivity and the presence of standardized assay formats, such as ELISA, latex agglutination and fluorescent polarization assays. Besides antibody, a large number of enzymes are also used directly as recognition molecules in IVD tests for metabolites and metal ion testing, such as glucose, lactate, K⁺ and Na⁺. These enzymes generate products and derivatives that can be quantified easily in the presence of the targets with high selectivity. Finally, as a result of recent advances in molecular diagnostics, nucleic acid probes (DNA or RNA) have been widely used to pinpoint the origins of various diseases and precise identification of infectious diseases through their highly specific nucleic acid hybridization. Although natural derived recognition molecules are widely used in IVD tests inside clinical laboratories, several limitations have limited their utility in the POC setting. For example, many antibodies and enzymes are generally considered to be less stable if they are not stored properly [29]; additionally, proteins purified from organisms often have large batch-to-batch variation, hence requiring careful calibrations, and limiting their use under POC settings.

Comparing to established affinity agents mentioned above, a range of new affinity agents that possess certain advantages have been discovered, isolated and tested for IVD applications as well as addressing some of the shortcomings of traditional antibodies and enzymes. For example, functional nucleic acids including aptamers and nucleic enzymes have been widely developed and tested as a substitute for antibodies and enzymes. they are widely regarded as more stable and economical replacement for antibodies as well as protein enzymes with a longer shelf-lives while possess the similar ability to recognize targets with high selectivity and activity. Additionally, since functional nucleic acids can be chemically synthesized, their batch-to-batch variation can be controlled more precisely. As a result, they have been used in numerous systems to develop IVD tests [30–35].

2.3. Signal transduction

Common signals to transduce target recognition include colorimetric, fluorescent, and electrochemical readouts. From the perspective of POCT, one of the main advantages of colorimetric modality is that it can potentially provide the detection results using the naked eye or little instrumentation [30]. For instance, lateral flow assays with colorimetric readouts are one example and have catalyzed the emergence of many commercialized products [30], such as pregnancy tests. This type of diagnostic modality is very attractive due to its cost-effective, easy-to-use, and rapid sample-to-answer, however, in most cases, it cannot provide quantitative information that is often indicative of disease severity. Additionally, fluorescent modality based on fluorescent labels, can provide more quantitative information [36], but often relies on dedicated instruments and is affected by fluorophore concentration, lamp intensity, and detector gain, making it difficult to calibrate, maintain and use [37]. One alternative approach is to utilize fluorescence polarization (FP) in fluorescence assays [37], because it is strongly sensitive to the rate of rotation of the fluorescent tag that depends on the target-induced structural or conformational change. More detailed information related to fluorescence polarization/anisotropy signaling mechanisms can be found in a recent review [38]. Another common modality is based on electrochemical techniques [39], which is being researched in many aspects of POCT because of its superiority for miniaturization, portability, and cost-effective. In addition, with the advance of nanotechnology and electrical engineering, electrochemical modality can offer ultrasensitive detection in native biological samples. As an example, Hajian et al. [40] developed a graphene-based field-effect transistor that uses clustered regularly interspaced short palindromic repeats (CRISPR) technology to enable the digital detection of a target sequence within intact genomic material. In this method, the sensor can detect the presence and concentrations of genomic DNA containing the target gene within 15 min by a simple handheld reader, with a sensitivity of 1.7 fM and without the need for amplification. Overall, each of the above-mentioned modalities has its own pros and cons, and further optimization is needed to reach the ASSURED criteria for effective POCTs.

2.4. Signal amplification

Signal amplification is another key element for IVDs, because the majority of the biomarkers are in low abundance in biological samples. One of the most common strategies is enzyme-catalyzed signal amplification using enzyme conjugates [39], which possesses several advantages such as high substrate specificity and rapid catalysis to produce electrochemical or optical signals. Enzyme-linked immunosorbent assay (ELISA) is one such example that has been widely used in clinical IVDs. Signal amplification through the integration of nanomaterials is another important tool used in various molecular assays [41]. In this approach, due to their high surface-to-volume ratio and size-dependent optical or electrochemical properties, the major function of nanomaterials is to serve as nanocarriers to immobilize numerous bioreceptors (e.g. antibodies and DNAs) to provide more binding sites for analytes, or integrate with numbers of optical/electrochemical signaling labels to produce enhanced signals. For instance, by integration of enzyme- and nanomaterials-based signal amplification, Ren et al. developed a dual-functional HRP-SiO₂@FFLuc NPs for in situ sequential detection and imaging of ATP and H₂O₂ in living mice [42]. Later, the same group reported fabrication of SiO₂ nanourchins through assembly of two-pronged pair of primers on mesoporous SiO₂ cores functionalized with Klenow DNA polymerase and DNA helicase [43]. Due to their high loading capacity, sufficient delivery of nucleic acid reagents into cells, efficient activity without any spatial mutual interference, and sustainable reactivity, the in situ amplified imaging of intracellular mRNAs in living cells was realized. Another new strategy of intracellular amplification-based dual-diagnosis and gene therapy were recently achieved through biodegradable nanosyringes (NSs), which were prepared by both covalent binding and electrostatic self-assembly of biomolecules onto free-standing silicon nanoneedles [44]. Recently, with the advance of DNA nanotechnology, DNA-based signal amplification strategies are emerging, and have demonstrated promising potentials for amplified biosensing for a broad spectrum of targets [45]. Some representative approaches include hybridization chain reaction (HCR), rolling circle amplification (RCA), nuclease-assisted target recycling, catalytic hairpin assembly (CHA) recycling, DNA nanomachines, and very recently CRISPR-based amplification. As a result of the discovery of collateral cleavage activity of several Cas proteins, a brand-new field is emerging and has revolutionized the IVDs for infectious diseases, with ultra-high sensitivity and selectivity. As an example, Chen et al. developed a method termed DNA endonuclease-targeted CRISPR trans reporter (DETECTR) by integrating Cas12a ssDNAse activation with isothermal amplification, which achieves attomolar sensitivity for DNA detection. DETECTR could enable rapid and specific detection of human papillomavirus in patient samples, thereby providing a simple platform for IVDs. Overall, the above-mentioned signal amplification strategies can also be combined together to produce more complex system to expand the scope of its application.

2.5. Multiplexing capability

Performing IVDs for multiple targets simultaneously from one single specimen is often clinically relevant, because measuring panels of biomarkers is expected to obtain more accurate diagnostic results than single biomarker [46]. To achieve a higher grade of multiplexing for IVDs, different methods have been developed [47–49]: (1) employing separate POCT devices or regional separating analytes into multiple detection zones by microfluidic chips, droplets, or arrays; (2) the use of different labels on different analytes, such as fluorescent dyes, enzymes, or nanomaterials; (3) the combination of these two strategies. As an example, Gootenberg et al. [49] reported a multiplexed and portable nucleic acid detection platform by screening out four distinguished Cas effectors with cutting preferences to different nucleotide sequences. By applying Cas13, Cas12a, and Csm6 in the CRISPR-based sensors, they further extended the detection to four targets by leveraging orthogonal dinucleotide motifs, with reporters for LwaCas13a, PsmCas13b, CcaCas13b, and AsCas12a in FAM, TEX, Cy5, and HEX channels, respectively. Despite the success, new strategies for multiplex IVDs are still limited, and thus further implementation with other emerging technologies is desirable. More importantly, although striving for multiplex diagnostics is important, it should be noted that a simple, sensitive, selective, fast detection system is far more important than a fancy but complex system for practical applications [47].

3. Advantages of using commercially available handheld meters for POCTs

Leading the list of the above newly developed IVDs technologies are those for point-of-care testing, which has been widely applied in many fields, such as personal healthcare monitoring in a hospital, clinic, or at home, on-site environmental assessment, and food safety analysis [50]. These tests generally consist of only elementary manual to use with little or no sample pre-treatment, and provide rapid response to small sample volumes [4]. Additionally, the commercially-available POCT readers range from transportable instruments, benchtop analyzers, to handheld devices or even portable pocket-size meters. To date, POC diagnostics have been comprehensively reviewed in the past decade [5, 51–53], from the development of new POC assays for diverse targets to the advancement in integration and miniaturization of POC devices.

The key criteria for such POC devices for IVDs are Affordable, Sensitive, Specific, User-friendly, Rapid and robust, Equipment-free and Deliverable to end-users (ASSURED) includes [16, 17]: (1) good usability, including simple and easy operation with eliminated or automated built-in maintenance functions; (2) sufficient accuracy, with simple or automated quality assessment; (3) accessibility for mobile healthcare and cloud-based data analysis; (4) multiplexing, that is simultaneous detection of different targets from a single sample. Overall, the emerging needs and demands for novel technologies for IVDs for diverse clinical applications is shifting the future of IVDs from central lab to point-of-care home testing.

Portable devices are growing in importance in applications of IVDs, however, despite many years of research, only a few portable meters are commercially available for non-specialists to make molecular measurements at home or in resource limited settings. A major challenge to develop such devices is the requirement of enormous financial and personnel expenses as well as time during the R&D process, making it difficult to be affordable to the general public. To address this issue, an alternative method is to integrate the well-developed molecular assays with current commercially available handheld meters through new signal transduction approaches. In this case, the developmental costs (e.g. time and financial) for a new POC device can be greatly decreased, while maintaining the advantages from both sides. Additionally, many existing portable devices have been marketed for decades (e.g., glucose meter, pH meter and thermometer), hence widely used and familiar to the general public. Directly make use of existing meters to the maximum extend can reduce the learning curve for any new POC IVD test significantly. Besides, by leveraging on existing platforms, any newly developed features can be easily introduced to the new POC IVD systems, such as wireless data transmission on some of today’s FDA approved smartphone enabled glucose meters. As a result, the adaptation of existing POC devices into a general measurement device via novel chemical or biological methods that can perform a wide range of common clinical tests could significantly increase the odds of bringing new POC IVD tests to reality.

4. Strategies to integrate molecular assays with commercially-available portable meters for POC testing

Some innovations in this field have been demonstrated to be versatile enough to support interchangeable biorecognition elements, ease of operation at a competitive cost, and remote detection of multiple, complex analytes. In the last decade, extensive signaling systems have been developed for quantitative analysis using commercially-available portable meters, such as personal glucose meter, pH meter, thermometer, and pressure meter. The advantages and disadvantages of these signaling systems are summarized in Table 1.

Table 1.

Advantages and disadvantages of various commercially-available portable meters adopted for POCT

Features	PGM	pH meter	Thermometer	Pressure meter
Wide availability	✔	✔	✔	✔
Low cost	✔		✔	✔
Easy-to-use	✔	✔	✔	✔
Real-time, continuous monitoring		✔	✔	✔
Small sample volume	✔		✔	✔
Resistance to interference from
Colored species	✔	✔	✔	✔
Autofluorescence	✔	✔	✔	✔
pH	✔		✔	✔
Glucose in biological matrix		✔	✔	✔
Temperature	✔	✔

4.1. Personal glucose meter

Personal glucose meter (PGM), firstly introduced around 1970, has become one of the most widely used POC devices that allows affordable detection of blood glucose levels [54]. With more than 40 years development, PGM displays unique features of low cost, portability, ease-of-use and rapid results, which benefits the diabetes around the globe. Despite the success, the common PGM can only detect blood glucose, which limits its application in molecular diagnosis. A major challenge in using a PGM as a general detector for non-glucose targets is to find a general method to link the presence and concentrations of other targets with a detectable glucose signal. From the perspective of the core element of a PGM, it is basically an amperometer that measures turnover of an electron-transfer mediator in the enzymatic oxidation of glucose, typically by glucose oxidase (GOx) or glucose dehydrogenase. Therefore, the detection of other targets is possible by replacing these glucose-responsive enzymes with other enzymes [55], such as lactate-responsive lactate dehydrogenase, cholesterol-responsive cholesterol oxidase and ethanol-responsive alcohol dehydrogenase. However, this method requires the change and modification of the PGM strips as well as the software of the PGM detector, thus limits its applicability in molecular diagnosis. An alternative method to expand the utility of PGMs to non-glucose targets was reported by Mogapatra et al. [56], where glucose was covalently linked to several enzyme substrates and subsequently applied for the detection of corresponding enzyme analytes. In this approach, the glucose can be released upon reaction of the substrate with the target enzyme, providing a rapid single-step assay for quantifying enzyme activities using a PGM. However, to apply this method, sophisticated synthesis of the enzyme substrates that covalently link to glucose is required. In addition, most of commercially-available PGMs are designed to monitor blood glucose, and thereby have a dynamic range of 0.6–33 mM for glucose, which is difficult for the detection of trace amount of biomarkers in blood. Therefore, a signal amplification method is necessary when integrating the molecular assays with a PGM. To meet the above challenges, three main signal transduction methodologies have been developed to repurpose the PGM for molecular diagnosis, including generation of glucose signal by enzymatic and non-enzymatic reactions, and generation of other PGM-detectable signals.

4.1.1. Generation of glucose signal by enzymatic reactions

The most common method to produce a PGM detectable signal is to use an enzyme to convert the PGM-inert substrate to the PGM-detectable glucose signal. For instance, in 2011, Xiang et al. reported breakthrough approaches to transform the selective binding of non-glucose targets with functional DNA-based receptors (e.g. aptamers, and DNAzymes) into a PGM-detectable glucose signal (Figure 2A) [10]. To this aim, invertase, which catalyzes the conversion of PGM-inert sucrose into glucose, was used as the signal transduction element, and conjugated with fDNAs via covalently bond. In the presence of a target, this fDNA-invertase conjugate is released into the solution due to a) the decrease of the melting temperature through the target-induced structure-switching of aptamers, or b) metal ion-dependent cleavage of DNAzyme substrate. In this case, the presence and concentrations of a target is correlated with the released amount of fDNA-invertase conjugate, and thereby is quantitatively related to the glucose generation by invertase-catalyzed reactions. More importantly, taking advantages of high enzymatic activity (k_cat = 1540 s⁻¹) of invertase [54], such a design allows the conversion of trace amount of targets into mM level of glucose that can be detected by commercially available PGMs. This concept has been further extended for the detection of many types of targets, including protein biomarkers [57–66], DNAs [67–76], miRNAs [77–79], DNA oxidative damage [80], small organic molecules [65, 69, 81–87], metal ions [88–91], enzyme activities [92–96], biotoxins [57, 97–101], pathogens [102, 103], and cancer cells [104].

Figure 2.

(A) Scheme of the method using a PGM to detect a wide range of targets beyond glucose. Reprinted from Xiang et al., 2011 [10] with permission of Nature Publishing Group. (B) Schematic illustration of detection process using glucose encapsulating liposome (GEL) on test trip device linked with a PGM. Reprinted from Zhao et al., 2017 [127] with permission of Elsevier. (C) Schematic illustration of NADH/PGM system for non-glucose target detection using NADH-dependent enzymes. Reprinted from Zhang et al., 2016 [129] with permission of Wiley. (D) Installing of logic gate on PGMs for the portable, resettable, and quantitative Point-of-Care detection of many targets in clinical care. Reprinted from Zhang et al., 2018 [130] with permission of Wiley.

Glucoamylase, or amyloglucosidase, which catalyzes the hydrolysis of PGM-inert amylose to glucose, is another alternative enzyme used in PGM-based sensors for non-glucose targets, including cocaine [105], ATP [105], hepatitis B surface antigen [106], DNA methyltransferase activity [107], miRNA [108, 109], microbial pathogen [110], thrombin [111], and C-reactive protein [111]. For instance, Yan et al. [105] developed a target-responsive “sweet” DNA hydrogel with entrapped glucoamylase for PGM-based portable and quantitative detection of non-glucose targets. In this approach, the presence of target molecule causes the disassembly of DNA hydrogel to release the entrapped glucoamylase, which subsequently catalyzes the hydrolysis of amylose to generate large amount of glucose for sensitive detection of the model target, cocaine and ATP.

Alkaline phosphatase (ALP), which catalyzes the hydrolysis of PGM-inert glucose-1-phosphate to glucose, has also been used to generate glucose signals for PGM-based sensors [112, 113]. For instance, Zhang et al. [112] have developed a simple method to link the enzymatic activity of galactose-1-phosphate uridyltransferase to the production of glucose, which allows POC diagnosis of galactosemia in authentic human clinical samples. Given the presence of ALP in numerous enzymatic assays for clinical diagnostics, this method can be adapted to quantify a wide range of enzyme activities using a PGM.

In addition to the above mentioned enzymes, other types of enzymes, such as β-galactosidase [114, 115], hexokinase [116, 117], and microbes-containing enzymes [118, 119], has also been integrated with PGMs for the development of portable sensors. Table 2 summarizes the analytical performance of some representative reports discussed above focusing on enzyme-based PGM sensors.

Table 2.

Analytical performance of some representative enzyme-based PGM sensors.

Enzyme Type	Real Samples	Analyte	Assay Format	Amplification method	Dynamic range	LOD	ref
Invertase	human serum	cocaine	sandwich	enzymatic	0–500 μM	3.4 μM	[10]
		adenosine			0–1,000 μM	18 μM
		IFN-γ			0–400 nM	2.6 nM
		UO22+			0–200 nM	9.1 nM
	human serum	streptavidin	sandwich	enzymatic	0–80 nM	4.0 nM	[57]
		PSA			0–100 ng/mL	1.5 ng/mL
		biotin	competitive		0–96 μM	1.5 μM
		OTA			0–1000 nM	17 nM
	human serum	CEA	sandwich	magnetic beads	0–1.0 ng/mL	0.05 ng/mL	[58]
	N.A.	insulin	sandwich	enzymatic	0–10 nM	1 nM	[59]
		HbA1c			0–200 mg/L	3 mg/L
	human serum	AFP	sandwich	enzymatic	0.5–50 ng/mL	0.18 ng/mL	[60]
	human serum	PDGF-BB	sandwich	CXR, CAMB	0.316 fM-1 pM	0.11 fM	[63]
	human serum	VEGF	sandwich	HCR	3–100 pg/mL	1.2 pg/mL	[64]
	human serum	8-OHdG	competitive	AuNPs	0.1–100 ng/mL	0.23 ng/mL	[65]
		PSA			1–100 ng/mL	1.26 ng/mL
	human serum	AFP	sandwich	BRCA	0.1–100 ng/mL	87 pg/mL	[66]
	N.A.	HBV DNA	sandwich	enzymatic	0–50 nM	40 pM	[67]
	N.A.	HIV DNA	sandwich	Fe3O4/Au NP	0.5 pM-1 nM	0.5 pM	[68]
	human serum	DNA	sandwich	ICSDPR	5.0–1000 fM	0.5 pM	[71]
	cell lysate	miRNA-21	competitive	HNFs	0.5–100 nM	0.41 nM	[78]
	milk	melamine	competitive	enzymatic	0–500 μM	0.53 μM	[82]
	FBS	amino acids	Label-free	enzymatic	0–20 μM	0.1 μM	[87]
	N.A.	Pb2+	competitive	enzymatic	0–1000 nM	16 nM	[88]
		UO22+			0–1000 nM	5.0 nM
	cancer cell	telomerase	sandwich	DNA machine	0.1 fM-1 nM	0.1 fM	[94]
	drinking water	MC-LR	sandwich	GO-AuNPs	0.60–100.00 ng/mL	0.10 ng/mL	[100]
	human serum	SW620 cell	sandwich	GO	1,560–100,000/mL	560 cells	[103]
Glucoamylase	urine	cocaine	label-free	hydrogel	0–750 μM	4.4 μM	[105]
		ATP			0–1 mM	N.A.
	human serum	DNA MTase	sandwich	liposome	0.005 to 0.1 U/mL	0.002 U/L	[107]
	cancer cells	miRNA-21	sandwich	P-ERCA	0.5–10 fM	0.1 fM	[109]
	lake water	E. coli	competitive	Fe3O4@SiO2	20–105 bacteria/mL	40 bacteria/mL	[110]
	human serum	thrombin	sandwich	liposome	0–200 nM	12.8 nM	[111]
		CRP			0–500 ng/mL	13 ng/mL
ALP	human blood	GALT	label-free	enzymatic	0–43.5 μmol h−1 (g Hb)−1	N.A.	[112]
Galactosidase	potable water	E. coli	label-free	enzymatic	20–2×109 CFU/mL	N.A.	[115]
Hexokinase	tap water, human serum, bovine urine	ATP	label-free	enzymatic	0.05–0.4 μM	49 nM	[116]

Abbreviations: IFN-γ = interferon-gamma; PSA = prostate-specific antigen; OTA = ochratoxin A; CEA = carcinoembryonic antigen. HbA1c = glycated hemoglobin; PDGF-BB = platelet-derived growth factor BB; CXR = cation exchange reaction CAMB = catalytic and molecular beacon; VEGF = vascular endothelial growth factor; 8-OHdG = 8-hydroxy-2’-deoxyguanosine; AuNPs = gold nanoparticles; BRCA = Backfilling rolling cycle amplification; HBV = hepatitis B virus; HIV = human immuno-deficiency virus; ICSDPR =isothermal circular strand-displacement polymerization reaction; HNFs = hybrid nanoflowers; FBS = fetal bovine serum; MC-LR = Microcystin-LR; GO-AuNPs = graphene oxide-gold nanoparticles; SW620 = human colorectal cancer cell; DNA MTase = DNA methyltransferase; P-ERCA = padlock exponential rolling circle amplification; ALP = Alkaline phosphatase; GALT = galactose-1-phosphate uridyltransferase; CRP = C-reactive protein; N.A. = Not Applicable; LOD = limit of detection;

4.1.2. Generation of glucose signal by non-enzymatic reactions

Instead of using enzymes to regulate the glucose signal for PGM-based sensing, another elegant method to generate a target-responsive glucose signal is to directly release encapsulated glucose from nanocontainers [120], such as mesoporous silica nanoparticles [121–125], liposome [126, 127], and TiO₂ nanotubes [128]. Compared with enzymatic generation of glucose signals, such a non-enzymatic controllable release system minimized the interferences from temperature and pH of solution, while maintains the advantages of easy operation and signal amplification. For instance, Fu et al. [122] reported the use of DNAzyme-capped mesoporous silica nanoparticles (MSNs) as glucose nanocontainers, and developed a PGM-based portable sensor for sensitive detection of lead ions. In this system, the presence of target Pb²⁺ ion catalyzes the cleavage of the DNAzymes, thereby opening the molecular gates and releasing encapsulated glucose from the MSNs. The released glucose molecules can be monitored by an external PGM, which is related to the presence and concentrations of the target Pb²⁺ ions. Using similar strategy, Gao et al. [123] designed a polystyrene microsphere-gated magnetic mesoporous silica nanocontainer for encapsulation of glucose, and combined with a competitive immunoassay for sensitive detection of brevetoxin B. Similar MSNs-based platform has also been applied for the detection of telomerase activity in cell extract using telomeric wrapping DNA as the gate.

Another non-enzymatic glucose generation system was recently developed by Zhao et al. [126], where antibody-tagged liposome was used as glucose container. The presence of target antigen leads to the capture of antibody-tagged liposome to form a sandwich complex, which can be lysed to release the encapsulated glucose for a PGM-based detection of protein biomarker. To simplify the detection process, the same group further developed an integrated disposable immunochromatographic strip with PGM readout for rapid quantification of phosphorylated proteins (Figure 2B) [127]. Later, TiO₂ nanotube arrays were also used as a nanocontainer and integrated with PGMs for portable and sensitive detection of cocaine [128]. Very recently, a 3D paper-based device was integrated with PGMs for highly sensitive and portable detection of silver ion [120]. The silver ion-triggered self-growing of Ag particles in the nanoporous membranes (NMs) of μPADs can partially block the pores of the NM, and as the solution flows through the NM and re-dissolves the glucose powder, rapid and efficient glucose signal generation can be achieved.

4.1.3. Generation of other PGM-detectable signals

When performing a molecular assay in blood or urine samples, the interference of endogenous glucose to PGM signals is problematic for many targets. To overcome this limitation, Zhang et al. [129] developed a new PGM-based signaling strategy based on its dose-dependent response to nicotinamide coenzymes (e.g. NADH) (Figure 2C). Taking advantage of this feature, a series of enzymatic reactions have been applied to link the target substrate to the consumption or production of NADH that could be monitored using a PGM. More importantly, an innovative strategy was developed to overcome the background glucose issue by employing an enzyme hexokinase that can efficiently convert the endogenous glucose to PGM-inert phosphorylated glucose during the PGM assay. As NADH is a functionally important cofactor associated with many clinical tests, this method can be readily expanded for the detection of a broad range of targets using various NADH-dependent enzymes. The invention of NADH-based signaling strategy is a milestone in the development of PGM-based practical sensors, allowing millions of people to use a single portable device to monitor not only their blood glucose levels, but also other relevant biomarkers.

Benefiting from the above glucose and NADH–based signal transduction strategies, a PGM-based biocomputing platform was recently developed by Zhang et al. [130], possessing logic capability for multi-parameter sensing of many clinically relevant targets (Figure 2D). In this approach, a series of metal ions, disease-related metabolites, coenzymes, and native enzymes have been applied as inputs, while glucose and/or NADH signal on a BGM was used as outputs. A complete set of seven binary logic gates (YES, NOT, OR, NOR, NAND, INHIBIT, and Concatenated) were constructed on PGMs for the monitoring of multiple biological substances, showing great promise for POC diagnostics of diseases, such as hyponatremia and hypernatremia. Significantly, this method will not only broaden the clinical applications of the BGM, but also open a new avenue in making PGM a logic-gate responsive POC device for practical applications.

In addition to the above PGM-based signaling strategies, other electroactive species, including paracetamol- or catechol-bearing compounds [131], reduced glutathione (GSH) [132], thiocholine [133], have also been demonstrated to generate a PGM-detectable signals for direct detection of β-galactosidase [131], α-mannosidase [131], influenza viruses [131], Streptococcus pneumoniae [131], and E. coli [131], intracellular glutathione and serum triglyceride levels [132], as well as organophosphorus pesticide [133].

In general, repurposing PGMs as a general measurement device may help in rapid practical translation of traditional diagnostic assays by bypassing costly device development process and sensor prototype validation.

4.2. pH meter

pH meter is another important type of commercialized portable meter, which measures the hydrogen ion activity in solutions. The obtained pH value indicates the degree of the acidity or alkalinity, with the unique features of pocket-size portability, low cost, easy operation and reliable high sensitivity (0.001 unit of pH change). As a result, numerous pH meters have been developed for diverse applications, such as healthcare and clinical assessment, water quality evaluation, and soil analysis. However, most of the commercialized pH meters are still limited to only detect a single “target”, pH. To reach the full potential of pH meters for molecular diagnostic applications, there is a need to find a method that can link the binding of any targets with a change of pH so any pH meter can be used to measure the presence and concentrations of targets of interests. To date, different signal transduction strategies based on pH meter have been developed, and can be categorized into two types, as detailed in the following sections.

4.2.1. Generation of pH change by enzymatic reactions

To generate a pH change of the assay solutions, the most used method is based on the catalytic reaction of bioactive enzymes that can produce the acid or alkaline species, such as glucose oxidase (GOx) [134], urease, and acetylcholinesterase (AchE) [135]. By applying these enzymes as labels and further integration with sandwich or competitive assay format (see section 2.2), the obtained sensors are able to transform the binding of target into a target-dependent pH change that can be detected using a pH meter. For example, Ye et al. [136] proposed a portable biosensor for on-site detection of a food pathogen through the combination of sandwich immunosensor and ubiquitous pH meter (Figure 3A). In this system, ConA-GOx-CaHPO₄ organic–inorganic hybrid nanoflowers were first prepared through a modified biomineralization process and applied as a signaling tag for Escherichia coli (E. coli) O157:H7 due to its high affinity to lipopolysaccharides O-antigen of E. coli. The readout signal was generated by the resulting pH decrease through the GOx-catalysis of glucose into gluconic acid after the sandwich immunoreaction. Using similar strategy, a series of pH meter-based portable biosensors have been developed for the detection of clinically relevant protein biomarkers, such as troponin I [135], human oncogenic protein [137, 138], vascular endothelial growth factor (VEGF) [134], carcinoembryonic antigen (CEA) [139], neuron-specific enolase (NSE) [140], and squamous cell carcinoma antigen (SCCA) [141]. In addition to the detection of proteins based on immunoreactions, Xie et al. [142] developed a pH meter based platform for the facile and sensitive detection of genomic DNA by directly measuring released hydrogen ions during the loop mediated isothermal amplification (LAMP) reaction catalyzed by DNA polymerase. Recently, to extend the targets to heavy metal ions, Tang et al. [143] designed a sensitive and portable pH meter based magnetic detection method for lead ion (Pb²⁺) by integration of Pb²⁺-specific DNAzyme, GOx with rolling circle amplification.

Figure 3.

(A) Illustration of the synthetic process for Con A-GOx-CaHPO₄ organic-inorganic hybrid nanoflowers and the corresponding scheme for immunoassay of E. coli O157:H7. Reprinted from Ye et al., 2016 [136] with permission of Wiley. (B) Schematic representation for the screening of thrombin inhibitors (lower four sensing areas) and detection of thrombin (upper four sensing areas) using a single eight-zone self-powered, portable, and light-addressable photoelectrochemical sensor. Reprinted from Wang et al., 2018 [147] with permission of ACS. (C) Translating molecular detections into a simple temperature test using a target-responsive smart thermometer. Reprinted from Zhang et al., 2018 [11] with permission of RSC. (D) Principle of the temperature-based immunosensor based on the exothermic reaction between H₂O and CaO using a common thermometer as readout. Reprinted from Ma et al., 2019 [151] with permission of ACS.

While the above sensors measure the decrease of the pH value in solution, urease, which catalyzes the hydrolysis of urea into ammonia that raises the pH value [12], has also been applied for the development of portable pH meter-based sensors [144]. For instance, taking advantage of a bacteria-specific RNA-cleaving DNAzyme probe as the molecular recognition element and the urease as a label, Tram et al. [12] translated the detection of bacteria into a pH increase, which can be readily detected using a litmus dye or pH paper. Recently, by replacing the DNAzymes with aptamers [144], antibodies [145], or telomerase specific primer [146], the portable detection of aflatoxin B1 in food samples or telomerase activity in cells have also been achieved.

Based on these studies, we summarize the analytical performance of some representative reports discussed above focusing on enzyme-based pH meter sensors, as shown in Table 3.

Table 3.

Analytical performance of some representative enzyme-based pH meter sensors.

Enzyme Type	Real Samples	Analyte	Assay Format	Amplification method	Dynamic range	LOD	ref
GOx	human serum	VEGF	sandwich	HCR	0.8–480 pg/mL	0.5 pg/mL	[134]
	human serum	HOP	sandwich	microparticles	0.620–40 pg/mL	0.57 pg/mL	[137]
	human serum	HOP	sandwich	microbeads	12.5–200 pg/mL	5.5 pg/mL	[138]
	human serum	CEA	sandwich	AuHM	0.1–100 ng/mL	0.062 ng/mL	[139]
	human serum	NSE	sandwich	liposomes	0.01–100 ng/mL	8.9 pg/mL	[140]
	human serum	SCCA	sandwich	HCR	0.01–10 ng/mL	5.7 pg/mL	[141]
	drinking water	Pb2+	label-free	RCA	1.0–100 nM	0.91 nM	[143]
AchE	human serum	troponin I	sandwich	enzymatic	0–100 ng/mL	10 pg/mL	[135]
DNA polymerase	silkworm egg	genomic DNA	label-free	LAMP	0.5 pg/μL-50 ng/μL	N.A.	[142]
Urease	Culture medium	bacterial	sandwich	enzymatic	5 – 5 × 105 cells	single CFU	[12]
	corn	AFB,	label-free	hydrogel	0.2–20 μM	0.1 μM	[144]
	corn	AFB,	sandwich	enzymatic	0.62–23.42 ng/mL	0.2 ng/mL	[145]
	human serum	telomerase	sandwich	enzymatic	50–10,000 cells	20 HeLa cells.	[146]

Abbreviations: GOx = glucose oxidase; VEGF = Vascular endothelial growth factor; HCR = hybridization chain reaction; HOP = human oncogenic protein; CEA = Carcinoembryonic antigen; AuHM = gold hollow microspheres; NSE = neuron-specific enolase; SCCA = squamous cell carcinoma antigen; RCA = rolling circle amplification; AchE = acetylcholinesterase; LAMP = Loop mediated isothermal amplification; N.A. = Not Applicable; LOD = limit of detection; CFU = Colony-Forming Units; AFB₁ = aflatoxin B₁;

4.2.2. Generation of pH change by non-enzymatic reactions

To measure the hydrogen ions concentration, the pH meter records voltage differences between two electrodes [147]. Based on this mechanism, Wang et al. [147] developed a portable, self-powered, and light-addressable photoelectrochemical (PEC) sensor for high-throughput screening of thrombin inhibitor drugs using a pH meter as signal readout (Figure 3B). The sensor was fabricated by immobilizing biotin-modified thrombin-cleavable peptides on eight separated sensing zones of a single gold film electrode, with a platinum (Pt) wire counter electrode that forms a self-powered PEC sensing platform. The presence of thrombin cleaves the biotin-modified peptides, leading to the decreased binding sites for streptavidin-labeled fullerene (C₆₀) PEC bioprobes. Due to the high conversion efficiency of C₆₀-based PEC bioprobe, the proposed thrombin sensor exhibits a wide detection range from 0.1 pM to 1.0 nM, with detection limit of 0.05 pM using a portable pH meter. In addition, the detection of thrombin and evaluation of inhibitor activity in serum samples have also been demonstrated.

4.3. Thermometer

The thermometer is one of the most commercial-available, easy-to-use and quantitative devices for portable measurements, which plays an important role in environmental monitoring and medical diagnostics. However, a major challenge in using a thermometer for the detection of other targets beyond temperature is to develop a method that can transform the binding of any targets into a temperature change so that the presence and concentration of a target can be detected using a thermometer. To address this issue, conventional calorimetric biosensors were developed [148], which utilize enzymatic reactions to generate heat and correlate it with the concentration of the enzyme substrate. However, these enzymatic reactions usually produce subtle change of temperature so that can only be detected using a highly sensitive and expensive instrument, which limits its application for POCT in resource-limited settings. As a result, the signal amplification is crucial for the successful development of thermometer-based portable sensors. To this aim, several exothermic reactions have been combined with the construction of thermometer-based biosensors in recent years for efficient signal amplification.

4.3.1. Generation of temperature signal by laser-assisted exothermic reaction

Photothermal effect is a common phenomenon associated with electromagnetic radiation, which is produced by the laser photoexcitation of a material, thereby producing thermal energy with an increase of temperature [149]. Using the laser-assisted exothermic reaction, Zhang et al. [11] translated molecular detections into a simple temperature test using a target-responsive smart thermometer (Figure 3C). The sensor system is based on the target-induced release of DNA–phospholipase A2 enzyme conjugate from a functional DNA duplex immobilized on magnetic beads, which subsequent catalyzes the hydrolysis of liposome-indocyanine green to produce a change of temperature signal under the NIR-laser irradiation. Considering the low cost and facile incorporation of the system with suitable functional DNAs to recognize many targets, the system makes the thermometer an affordable and pocket-size meter for the detection and quantification of a wide range of targets.

4.3.2. Generation of temperature signal by non-laser exothermic reaction

In addition to the laser-assisted exothermic reaction, the dissolution of some inorganic oxide in water can also generate a large amount of heat to change the solution temperature. Based on this phenomenon, Gao et al. [150] reported an exothermic chip that could generate a temperature change based on a target-responsive aptamer-modified hydrogel. The presence of an analyte could increase the capillary flow rate for the acceleration of NaOH dissolution in water, resulting in a quick increase of temperature that can be measured using a forehead thermometer. Using this method, heavy metal ions (Hg²⁺ and Pb²⁺) in different real samples are quantitatively analyzed. Recently, Ma et al. [151] further designed a temperature-based immunosensor (Figure 3D), which transduces the antibody–antigen recognition event into the temperature change with a commercialized thermometer as signal readout. By taking advantages of high catalysis efficiency of nanozyme and self-heating reaction between CaO and H₂O, the quantitative detection of CEA can be realized without the assistant of any expensive instruments or laser sources other than a simple thermometer.

4.4. Pressure Meter

Pressure meter is also a common portable device for measuring pressure generated by gases or liquids during many chemical or biochemical reactions. With the advantages of high sensitivity and portability, pressure meter is emerging as a promising candidate for POC diagnostics. Yang’s group pioneered the development of pressure-based biosensors by translating the molecular recognition signal into a measurable pressure signal in a sealed device through a gas-generation reaction. For example, Zhu et al. designed pressure-based bioassays by integrating a sandwich immunoassay with a common gas-generation reaction catalyzed by a catalase or Pt nanoparticles [152]. The presence of a target results in the formation of the sandwich immunocomplex that triggers the decomposition of substrate H₂O₂ to H₂O and O₂, leading to significant pressure increase in the reaction chamber that can be measured using a pressure meter. Taking advantages of the high liquid to gas volume expansion and the excellent catalytic properties of catalase/PtNP, this pressure-based method was able to provide more than 10¹⁰ folds of signal enhancement with a few minutes of reaction. Taking advantage of this feature, Yang and other groups has developed a series of pressure meter-based biosensors for the detection of diverse targets, including PSA, Influenza A/H5N1 virus (H5N1) [152], C-reactive protein (CRP) [153], cocaine, Pb²⁺, OTA [154], cancer cells [155, 156], telomerase activity [157], myoglobin [158, 159], miR-21 [160], Hg²⁺ [161], and pathogenic bacteria [162].

4.5. Other meters

In addition to the above mentioned portable sensors that utilize glucose, pH or temperature as the signal readout, some important metabolites, such as ATP [163, 164], uric acid [165], have also been applied as the signal readout that can be quantified using a commercial-available portable meter. For instance, Chen et al. [164] integrated the double-enzymes-mediated bioluminescent reaction, conjugated nanoparticles, and portable ATP detector to construct a rapid, highly sensitive, and quantitative POCT assay for proteins. The proposed sensor possesses the features of high sensitivity (femtomolar level of biomarkers), fast sample-in-answer-out (< 1h), and convenient operation (portable ATP detector), which broadens the range of portable sensors in molecular diagnosis.

5. Conclusion and perspectives

In summary, we have witnessed a rapid growth in developing IVDs in the past decade. At the fundamental level, we briefly discussed the typical workflow for IVDs from sample preparation to target recognition, signal transduction, signal amplification and multiplex capability, and highlighted several new innovative technologies in each element of the workflow. Owing to the demand for personalized medicine, IVDs is shifting from centralized lab to POCT. POC IVDs developed for resource-limited settings is becoming widespread acceptance for doctor’s office, clinic, or home use, and these POCTs will play an important role in reducing medical costs for everyone. From the perspective of product development, the adaptation of existing POCT devices for detection and management of new targets can significantly increase the odds of bringing new POC tests to reality. To date, various commercially-available portable meters have been repurposed to broad their utility in POC IVDs through the integration of many innovative signal transduction strategies.

Despite the significant progress, several issues related to current and future practical applications in POC IVDs still need to be addressed. The first issue is reagents storage and release in POC devices. To integrate molecular assays with existing POC devices for home monitoring, it is necessary to have all necessary sensor reagents pre-stored in testing strips or cartridges of POC devices. Some microfluidic chips have been developed for reagents storage, which could to some extent eliminate the manual reagents introduction, simplify the detection process, and minimize the contamination risk, making POC devices much friendly to end-users. Deng et al. [166] recently provided an excellent and timely overview of the advances in reagents storage and release in self-contained POC devices. However, the reagents involved in POC testing vary widely, and thus each of the storage and release strategy requires further optimization for new conditions. Future advances in microfluidic technologies and engineering strategies, as well as the creation of new stable reagents, and innovation in nanotechnology, will provide more opportunities tackle the limitations in this aspect.

Another consideration from a POC perspective for home use is user competency, which is one of potential source of error in POC tests. Compared with molecular testing in centralized lab, POC testing by patients may have higher error rates due to the lack of training during sample collection and analysis. It is therefore important to develop more automatic platforms for POC diagnostics, because a high level of automation in sampling process will facilitate the generation of accurate results for POC tests at home. Current existing portable meters, such as PGMs, have significantly improved analytical performance, and have been engineered to eliminate many of the potential error-generating steps in the testing process.

Finally, the connectivity is another important aspect in current and future POC diagnostics, which is vital for effective management of testing results and quality assurance, particularly in the new era of telemedicine. With the advance of mobile technology, mobile POC devices are emerging to broaden the current POC diagnostic landscape. A broad collaboration of experts in chemistry, engineering, and computer science is desired to push the community toward the development of more effective POC devices.

Highlights.

• Typical workflow for in vitro diagnostics and innovative technologies to improve the workflow are summarized.

• Advantages of using commercially available handheld meters for POCT are highlighted.

• Comprehensive coverage of signal transduction strategies to repurpose the commercially-available handheld meters for POCT are presented.

• Future challenges and prospective of the applying POCT are discussed.

List of Abbreviations:

POC

Point-of-care

IVD

In vitro diagnostics

PCR

Polymerase chain reaction

LOC

Lab-on-a-chip

LID

Lab-in-a-drop

μPADs

Microfluidic paper-based analytical devices

CRISPR

Clustered regularly interspaced short palindromic repeats

ASSURED

Affordable, Sensitive, Specific, User-friendly, Rapid and robust, Equipment-free and Deliverable to end-users

IFN-γ

Interferon-gamma

ELISA

Enzyme-linked immunosorbent assay

CHA

Catalytic hairpin assembly

DETECTR

DNA endonuclease-targeted CRISPR trans reporter

R&D

Research and development

PGM

Personal glucose meter

MSNs

Mesoporous silica nanoparticles

NMs

Nanoporous membranes

NADH

Nicotinamide adenine dinucleotide

GSH

reduced glutathione

PSA

prostate-specific antigen

OTA

Ochratoxin A

CEA

Carcinoembryonic antigen

HbA1c

Glycated hemoglobin

PDGF-BB

Platelet-derived growth factor BB

CXR

Cation exchange reaction

CAMB

Catalytic and molecular beacon

VEGF

Vascular endothelial growth factor

8-OHdG

8-hydroxy-2’-deoxyguanosine

AuNPs

Gold nanoparticles

BRCA

Backfilling rolling cycle amplification

HBV

Hepatitis B virus

HIV

Human immuno-deficiency virus

ICSDPR

Isothermal circular strand-displacement polymerization reaction

HNFs

Hybrid nanoflowers

FBS

Fetal bovine serum

MC-LR

Microcystin-LR

GO-AuNPs

Graphene oxide-gold nanoparticles

SW620

Human colorectal cancer cell

DNA MTase

DNA methyltransferase

P-ERCA

Padlock exponential rolling circle amplification

ALP

Alkaline phosphatase

GALT

Galactose-1-phosphate uridyltransferase

CRP

C-reactive protein

N.A.

Not Applicable

LOD

Limit of detection

ConA

Concanavalin A

GOx

Glucose oxidase

HCR

Hybridization chain reaction

HOP

Human oncogenic protein

CEA

Carcinoembryonic antigen

AuHM

Gold hollow microspheres

NSE

Neuron-specific enolase

SCCA

Squamous cell carcinoma antigen

RCA

Rolling circle amplification

AchE

Acetylcholinesterase

LAMP

Loop mediated isothermal amplification

CFU

Colony-Forming Units

AFB₁

Aflatoxin B₁

PEC

Photoelectrochemical