Point-of-Care Diagnostics Using Self-heating Elements from Smart Food Packaging: Moving Towards Instrument-Free Nucleic Acid-Based Detection

Abstract

Compromising between accuracy and rapidity is an important issue in analytics and diagnostics, often preventing timely and appropriate reactions to disease. This issue is particularly critical for infectious diseases, where reliable and rapid diagnosis is crucial for effective treatment and easier containment, thereby reducing economic and societal impacts. Diagnostic technologies are vital in disease modeling, tracking, treatment decision making, and epidemic containment. At the point-of-care level in modern healthcare, accurate diagnostics, especially those involving genetic-level analysis and nucleic acid amplification techniques, are still needed. However, implementing these techniques in remote or non-laboratory settings poses challenges because of the need for trained personnel and specialized equipment, as all nucleic acid-based diagnostic techniques, such as polymerase chain reaction and isothermal nucleic acid amplification, require temperature cycling or elevated and stabilized temperatures. However, in smart food packaging, there are approved and commercially available methods that use temperature regulation to enable autonomous heat generation without external sources, such as chemical heaters with phase change materials. These approaches could be applied in diagnostics, facilitating point-of-care, electricity-free molecular diagnostics, especially with nucleic acid-based detection methods such as isothermal nucleic acid amplification. In this review, we explore the potential interplay between self-heating elements, isothermal nucleic acid amplification techniques, and phase change materials. This paves the way for the development of truly portable, electricity-free, point-of-care diagnostic tools, particularly advantageous for on-site detection in resource-limited remote settings and for home use.

Key Points

Electricity-free nucleic acid-based detection can be achieved using self-heating elements adapted from smart food packaging technology.

The integration of self-heating elements and phase change materials with isothermal nucleic acid amplification techniques offers a promising solution for developing portable electricity-free diagnostic devices at the point-of-care level, and in this review, examples of such devices for pathogen detection were examined.

Incorporating self-heating technologies, already approved for commercial use in food packaging, into diagnostic tools could revolutionize healthcare by enabling precise genetic-level diagnostics, which are particularly valuable during outbreaks and in resource-limited areas.

Introduction

The coronavirus disease 2019 pandemic has highlighted the critical need for rapid and reliable diagnostic tools, particularly for the early detection of infectious diseases. The lack of prompt and accurate diagnostics in the early stages of the pandemic contributed to the rapid global spread of severe acute respiratory syndrome coronavirus 2 virus in 2019 [1]. Diagnostic accuracy is essential not only for treatment but also for controlling the transmission of diseases, a point emphasized by global health organizations such as the Centers for Disease Control and Prevention, European Centre for Disease Prevention and Control, and the World Health Organization [2–4].

While nucleic acid amplification methods like polymerase chain reaction (PCR) tests remain the gold standard in detection because of their high sensitivity and specificity, they require specialized equipment and trained personnel [2, 5]. This makes PCR tests costly, time consuming and less accessible, particularly in resource-limited settings. Antigen tests, although faster and more convenient to use outside the laboratory, typically offer low sensitivity, due to factors such as mutations and individual immunity [6]. This gap in rapid, accurate, and accessible diagnostics underscores the need for point-of-care testing (POCT) solutions that can be effectively utilized outside traditional laboratory settings [7].

One of the primary challenges in developing portable nucleic acid-based POCT lies in the need for precise temperature control, especially in techniques such as PCR, which requires repeated thermocycling. Isothermal amplification methods, developed over the past two decades, offer a promising alternative by minimizing the need for complex temperature cycling, though they still require controlled (elevated) temperatures.

To overcome these challenges in non-laboratory settings, incorporating microheaters and microsensors, either independently or within microfluidic systems, presents a practical and effective approach. Microheaters generate heat through mechanisms such as Joule heating, ultrasonic heating, or radiative methods, are regulated by electrical, mechanical, and thermal properties, as well as its material composition and geometric design [8]. When integrated into microfluidic platforms, microheaters enable the development of portable diagnostic kits for on-site detection, thereby reducing dependence on external laboratory infrastructure [9]. Additionally, the integration of microsensors, particularly those utilizing electrical impedance analysis, significantly enhances diagnostic capabilities by enabling rapid and accurate nucleic acid detection. These microsensors, fabricated using microelectromechanical systems technology [9, 10]. However, despite their potential, microheaters and microsensors present certain drawbacks, including power consumption, uneven heat distribution, material limitations affecting long-term reliability, and high fabrication costs associated with microelectromechanical system technology, which can limit their accessibility [9].

In contrast, a promising and innovative solution is the integration of self-heating elements with phase change materials (PCMs) for temperature regulation. These technologies, already employed in smart food packaging, could be adapted for diagnostic applications, presenting a pathway to portable electricity-free diagnostic tools.

In this outlook review, we first explore the importance of POCT and nucleic acid-based amplification in diagnostics. We then provide an overview of self-heating elements used in smart food packaging. Finally, we discuss how self-heating elements from the food industry can integrate with nucleic acid amplification to create a portable device, point-of-care (POC) solution for prompt and accurate diagnosis.

Point-Of-Care Testing

Point-of-care testing refers to diagnostic testing conducted outside a central hospital laboratory, typically near the patient’s care site, such as at the bedside, in a physician’s office, or in the operating room [11–14]. Point-of-care testing enables immediate testing and analysis of patient samples, allowing for rapid diagnostic to assist caregivers in making timely treatment decisions [11, 13, 15]. Point-of-care testing can be deployed in various testing settings, including emergency departments, critical care units, outpatient clinics, and even non-traditional environments such as schools, workplaces, and mobile nursing practices [13]. The key advantages of POCT include increased accessibility to diagnosis, minimal sample volumes, and rapid analysis times, with the ideal goal of providing test results within minutes using simple protocols [15]. However, a single parameter POCT is often more expensive compared with the clinical laboratory diagnostics. The cost effectiveness of laboratory diagnostics continues to be the main hurdle for extensive use of POCT in many cases, especially in countries with well-developed laboratory infrastructures [16].

In addition to metabolites such as glucose, lactate, and a few other small molecules, POC diagnostics can be classified into two main groups: protein testing and nucleic acid testing [17, 18]. These two crucial pillars in diagnostic medicine offer distinct advantages and applications. Point-of-care protein testing, such as immunoassays characterized by its rapidity, cost effectiveness, and simplicity, has become indispensable tools [19–21]. For example, during the coronavirus disease 2019 outbreak, rapid testing became a frequently used tool for controlling the pandemic, with at-home antigen tests widely available to the public, though less accurate compared with nucleic acid testing [2].

Microfluidic chips and paper-based devices are at the forefront of POC development [22–25]. While microfluidic chips offer high sensitivity and specificity, their requirement for additional equipment complicates their implementation. Conversely, paper-based assays, with their affordability and ease of use, hold significant promise for widespread adoption. Recent advancements in microfluidics and nanotechnologies have propelled the rapid evolution of paper-based POC assays, with commercially available products such as lateral flow immunoassays showcasing their potential in diagnostics [23, 26].

Analysis based on nucleic acids offers significant advantages for diagnosing various conditions, including infectious diseases, tumor biomarkers, and drug metabolism [27]. Point-of-care testing of nucleic acids enables direct pathogen detection without the need for culturing, providing speed and convenience [27, 28]. Consequently, research and development efforts are focused on microfluidic-based POCT platforms that integrate sample preparation, nucleic acid extraction, amplification, and detection within a single device, enabling rapid and sensitive detection of DNA and RNA targets [18, 29]. Despite challenges related to affordability and accessibility, ongoing advancements in nanotechnology hold promise for further enhancing nucleic acid POCT and expanding its clinical applications [28].

While POCT is becoming increasingly available in clinical settings, home-based POCT offers even more applications and convenience for customers to use in the comfort of their homes [30]. These at-home diagnostic tools enable individuals to monitor their health and make informed decisions about their care [13, 30]. For example, home pregnancy tests have become a widely accepted practice [31]. Similarly, glucose monitoring for diabetes mellitus management has advanced significantly, allowing patients to manage their disease more effectively through regular self-testing with handheld glucose meters [30, 32, 33]. Beyond these, the range of available tests has expanded to include assessments for infections such as HIV and sexually transmitted diseases, and chronic conditions such as cardiovascular disease and osteoporosis [13, 31]. Technological advancements, such as telemedicine and telehealth, further enhance the effectiveness of home testing by enabling remote data transmission and allowing healthcare professionals to monitor patients’ health status from a distance [34]. The growing trend towards proactive healthcare management and personalized medicine is expected to continue expanding as technology advances and regulatory frameworks adapt, ultimately empowering individuals to take control of their health and well-being [13, 14].

Despite the growing potential of POC home testing diagnostics, several challenges persist. These tests must be designed for ease of use by individuals without laboratory training, with simple and intuitive procedures to ensure accurate results [35]. Additionally, the single-use nature of POC tests can be both inconvenient and costly compared with bulk testing in centralized laboratories [36]. Furthermore, POC test kits need to be resilient to environmental factors such as light, humidity, and temperature fluctuations, as poor storage conditions can compromise their accuracy [35].

Nucleic Acid Amplification in Diagnostics

Since the discovery of the DNA double helix structure in the 1950s [37], molecular diagnostics have evolved from a scientific tool into a routine part of medical care (Fig. 1). Among these advancements is Kary Mullis’ pioneering invention of the PCR in the 1980s [38, 39], which marked a significant shift in molecular diagnostics. Polymerase chain reaction revolutionized this field by lowering detection thresholds to trace amounts and even to single molecules, thanks to its ability to amplify DNA segments. Recognized for its exceptional sensitivity and selectivity, PCR became an instrumental tool in mutation exploration and the diagnosing of diverse pathologies [40]. This method involves cycles of heating and cooling DNA for repeated denaturation, primer annealing, and extension. The process begins with denaturation of the double stranded DNA at 93–95 °C. The annealing temperature is then determined by the melting temperature of the selected primers for the specific target. Finally, the primer extension step is carried out at 72 °C. By employing a thermostable DNA polymerase enzyme and specific primers, this method can achieve amplification exceeding a million-fold under an hour [38, 39, 41–45]. However, in order to achieve a successful amplification, specialized thermal cycling instruments are essential to this technique.

Fig. 1.

Milestones of molecular diagnosis techniques and self-heating elements. LAMP loop-mediated isothermal amplification, PCR polymerase chain reaction, RPA recombinase polymerase amplification

With the aim of amplifying nucleic acids without the need for thermal cycling, isothermal nucleic acid amplification (INAA) methods were developed [46–48]. isothermal nucleic acid amplification procedures operate at a single reaction temperature, using a special DNA polymerase with strong strand displacement activity and lacking 5′→3′ and 3′→5′ exonuclease activity, which significantly simplifies operational protocols [49]. The rise of INAA methods as viable alternatives to PCR began in the early 1990s, moving away from PCR’s reliance on sophisticated temperature cycling for denaturation, annealing, and extension [46, 47].

A range of INAA methods includes primer-generation rolling circle amplification [50, 51], recombinase polymerase amplification (RPA) [52], helicase-dependent amplification [53], strand displacement amplification [46], loop-mediated isothermal amplification (LAMP) [47], cross-priming amplification [54], exponential amplification reaction [55], whole genome amplification [56], and isothermal multiple-self-matching-initiated amplification [57]. Each method utilizes a different number of primers and enzymes, and some may require additional proteins, functioning at specific reaction temperatures ranging from physiological temperature up to 65 °C. In recent years, RPA and LAMP have been most prominently investigated for POCT applications, often combined with visual readout methods such as pH-dependent color change or lateral flow assays [58].

All INAA methods still require temperature control, especially at elevated temperatures: 37–42°C for RPA or 60–65°C for LAMP, for example. Therefore, a source of heating and temperature control must be provided at the point of need, and most proposed solutions involve electronic devices for heating and control. However, a truly independent POCT device should function without reliance on any instrument. The ideal POCT device is self-contained, which is where technological developments from other fields come into play.

Self-heating Elements in Smart Food Packaging

Beyond diagnostic developments, the food industry has begun designing self-contained systems for warming and cooling food and beverages in outdoor settings since the 1950s (Fig. 1). In fact, as early as the late 19th century, methods to heat and cool food and beverages without relying on external sources were already being investigated [59, 60]. A significant milestone in the development of self-heating packaging occurred in late 20th century with the introduction of flameless ration heaters, which utilize exothermic reaction processes [61]. Initially developed for and used by the US Army, flameless ration heaters are now available to consumers for heating “Meal, Ready-to-Eat” products [62, 63]. This method uses an exothermic reaction, typically water and calcium oxide (quicklime), to generate heat in a controlled and safe manner [62, 64]. Exothermic reactions are chemical processes that release energy, such as heat, into their surroundings. A common example is the hydration of quicklime (calcium oxide). When quicklime is combined with water, an exothermic reaction occurs, producing calcium hydroxide (a hydrate) and releasing thermal energy. This reaction causes the mixture to expand or “puff up.” The calcium hydroxide can be reconverted into quicklime by heating it to high temperatures, removing the water and reversing the hydration reaction. The amount of energy released during an exothermic reaction depends on the quantity of water involved and the nature of the reaction itself [65].

For example, a self-heating commercialized beverage from “the 42 Degrees Company” can increase its temperature by + 42 °C in just 3 min (Table 1). This rapid heating is achieved through the exothermic reaction between quicklime and water, which takes place in a compartment within the beverage can. The can is designed with two separate compartments: one contains water and calcium oxide, while the other holds the beverage. These compartments are completely isolated, ensuring that the chemical reaction never comes into direct contact with the drink. The process begins when the bottom of the can is pressed, breaking a seal that initiates the reaction by mixing water to calcium oxide. The heat generated is then safely transferred to the beverage, efficiently raising its temperature for convenient consumption.

Table 1.

Current applications of exothermic heat reactions in commercially available smart food packaging products on the market

Exothermic reaction	Product	Companies using exothermic heat reactions
Calcium oxide (CaO) + H2O	Hot beverages, soups, and foods	Aventura Self Heating (https://aventuradrinks.com/) The 42 Degrees Company (https://www.the42degreescompany.com/) HotCan (http://hotcan.com/)
Magnesium-based reaction (Mg-Fe) + H2O + salt	Hot beverages and meals	HeaterMeals (https://heatermeals.com/) OMEALS (https://www.omeals.com/) (https://www.isellpackaging.com/) Henan Wordgen International Co. Ltd (https://xxwordgen.com/product-page/)
Aluminum powder (AI) + calcium oxide (CaO) + sodium bicarbonate (NaHCO3) + H2O	Heating food and drinks	http://www.powdermaterial.com/product/self-heating-package-for-food
Aluminum powder (AI) + calcium oxide (CaO) + H2O	Heating food and drinks	Henan Wordgen International co. ltd (https://xxwordgen.com/product-page/)
Aluminum and silica	Hot beverages	HeatGen (https://www.poweredbyheatgen.com/)

The evolution of self-heating solutions reflects a pursuit of convenience, efficiency, and versatility in food packaging. Today’s sophisticated systems owe their development to technological breakthroughs, safety considerations, and consumer demand [66]. Table 1 showcases current commercially available smart food packaging products incorporating exothermic heat reactions, which utilize four specific chemical reactions.

Combining Self-heating Elements and Nucleic Acid Amplification for POC Diagnostics

Instrument-Free Nucleic Acid Amplification Devices Based on Exothermic Reactions with Water

In this section, diagnostic devices that successfully incorporated concepts and commercially available exothermic reactions from the food packaging industry are reviewed (Table 2, Fig. 3). In 2008, Weigl et al. proposed non-instrumented and/or minimally instrumented diagnostic devices that generate heat through a calcium oxide reaction and regulate temperature with a PCM, such as paraffin, making them suitable for INAA methods such as LAMP [67]. The first instrument-free device based on INAA and an exothermic reaction was the Non-instrumented Nucleic Acid Amplification (NINA) platform, developed by LaBarre et al. in 2009 [68, 69] (Fig. 3a). This platform aimed to provide a reliable test for diagnosing infectious diseases in a low-resource setting by utilizing the calcium oxide exothermic reaction. Once heat is generated by adding water to the exothermic reaction—using calcium oxide in this work—temperature stabilization in NINA devices is achieved by surrounding the amplification chamber with a layer of PCMs (Fig. 3a). These PCMs absorb the heat energy generated by the exothermic reaction and store it as latent heat for extended periods, effectively maintaining an isothermal temperature [70, 71].

Table 2.

Instrument-free, electricity-free, nucleic acid amplification devices based on exothermic reactions and water

Sample format, pathogen	Detection time	Nucleic acid amplification method	Water-base exothermic reaction	Type of PCMs	Readout	Limit of detection/sensitivity	Publication year	Refs.
Purified DNA of a bacteria (Ralstonia solanacearum race 3 biovar 2)	~ 60 min	LAMP	CaO	PureTemp 65a	Fluorescence	137 genome copies	2011	[70]
Plasmodium falciparum genomic DNA	~45 min	LAMP	CaO	Engineered phase change materials	Fluorescence	~50 parasite genomes/μL	2011	[71]
Escherichia coli DNA	~ 60 min	LAMP	Mg-Fe + NaCl in a microfluidic polymeric cartridge format	Paraffin frame	Fluorescence	10 target molecules	2011	[82]
Purified RNA and DNA spiked into human plasma, HIV-1	Not mentioned	RT-LAMP/LAMP	CaO	With PCM not mentioned which type of PCM	Fluorescence	For HIV-1 DNA: 10 copies per reaction/for HIV-1 RNA: 140 copies per reaction	2012	[74]
DNA isolated from a bacteria, Salmonella enterica-cell culture serial dilutions in 2% milk purified by SNAP card	~ 3 h including sample preparation	LAMP	CaO	PureTemp 65	Fluorescence	1 CFU/mL	2013	[81]
HIV-1 DNA	20–30 min	RPA	Sodium acetate trihydrate (SAT)	Without PCM	Immunochromatogra phic strip (ICS)	10 copies	2014	[83]
Malaria Plasmodium genus	~ 60 min	LAMP	Mg–Fe pouch format	With PCM not mentioned which type of PCM	Turbidity	The sensitivity is 96.8% (95% CI 83.2–99.5)	2015	[93]
Malaria P. falciparum parasites						The sensitivity is 100% (95% CI 75.1–100)
Saliva samples spiked with Zika virus	Less than 40 min	RT-LAMP	Mg-Fe Pouch format	PureTemp 68	Naked-eye Colorimetric via microfluidic cassette	Sensitivity: 5 PFU	2016	[84]
Influenza A (H1N1) virus	35 min	PCR	CaO	Water	Naked-eye lateral flow strip	1.0 TCID50/mL	2017	[85]
Human papillomavirus DNA	~ 70 min	LAMP	Mg-Fe alloy pouch format	PureTemp 68	Colorimetric via microfluidic chip	50 copies per test	2019	[73]
DNA extracted from T4 bacteriophage	~10 min	RPA	Lithium	Without PCM	Fluorescent assay	Not mentioned	2020	[86]
DNA isolated from Cronobacter species bacteria	Less than 2 h and 30 min	LAMP	CaO	Three types of PCMs: suitable for maintaining temperatures at 35 °C, 65 °C, and 100 °C	Naked-eye Colorimetric Visual detection by hydroxynaphthol blue	102 cfu/g	2020	[94]
saliva samples containing adenovirus DNA	More than an hour	LAMP	CaO + Al	Palmitic acid	Visual detection by color change	2 copies/μL	2023	[95]
Escherichia coli DNA	30 min	RPA	CaO + sodium carbonate	Without PCM	Lateral flow strip	36.23 CFU/mL	2024	[87]

CFU colony-forming unit, CI confidence interval, h hours, min minutes, PCM phase change material, PFU plaque-forming units, Ref reference, RPA recombinase polymerase amplification, RT-LAMP reverse transcriptase-loop-mediated isothermal amplification, TCID tissue culture infective dose, TCID50 the quantity of viruses required to cause cytopathic effect with half of cells in a cell culture plate

^aWith a melting point of 65 °C

Fig. 3.

Schematics of different water-activated exothermic reaction instrument-free, electricity-free nucleic acid amplification devices. a The first version of the Non-instrumented Nucleic Acid Amplification (NINA) device [69]. b Self-heating non-instrumented cartridge for isothermal amplification of nucleic acids [82]. c An instrument-free device based on a capillary tube for convective polymerase chain reaction thermocycling, utilizing a calcium oxide mixture and lateral flow detection [85]. d Schematic of miniature lithium heaters with water [86]. PCM phase change material, POM Polyoxymethylene, SDS sodium dodecyl sulfate

In different versions of NINA devices, two types of exothermic reactions are used for heat generation: calcium oxide and magnesium-iron alloy [70–72]. The combination of exothermic reaction with PCMs for heat generation is a key development for the NINA devices. Phase change materials are compounds vital for managing thermal energy through storing energy as latent heat by inhibiting a phase transition [70, 71, 73]. They function optimally at specific melting temperatures, utilizing latent heat to maintain temperature stability within a defined range. Phase change materials find applications in diverse areas, such as energy management in buildings, solar energy storage, textiles, and smart food packaging [75–77]. Phase change materials, known for their adeptness in manipulating thermal energy during phase transitions, are particularly advantageous in handling the substantial heat generated during exothermic reactions [78, 79]. In the context of an exothermic reaction, the heat released initiates the melting process of the PCM, with the additional heat absorbed as latent heat of fusion within the PCM [80]. Once equilibrium is achieved, the stored latent heat sustains the two-phase PCM at the target temperature until the solid-to-liquid transition is completed and it solidifies entirely [75] (Fig. 2). The chosen PCM in NINA devices stands out for its adjustable characteristics, including the melting temperature range, specific heat capacity, and thermal conductivity, rendering the device a versatile incubation platform suitable for specific isothermal amplification techniques [71].

Fig. 2.

Thermal energy stored in phase change materials (PCMs). The graph illustrates the relationship between temperature (T) and thermal energy (Q) for a PCM, indicating the energy absorbed by the PCM during the phase change at constant temperature, providing insight into PCM heat storage dynamics

Since 2009, multiple iterations of the NINA platform have been successfully developed, with each version designed to detect a range of pathogens (Table 2) [70, 71, 74, 81]. In 2015, Diesburg et al. undertook a redesign of the NINA device with the aim of transforming it into a single-use disposable platform referred to as NINA-SUD. This new design approach involved re-engineering PCMs, by incorporating thermal enhancement additives and implementing a controlled introduction of reactants through porous media [72].

In addition to these NINA device versions, there are other devices for nucleic acid amplification techniques that utilize exothermic reactions with water as a heat source. These include the following devices.

In 2011, Liu et al. pioneered the development of a disposable, water-activated, self-heating, non-instrumented microfluidic cartridge for INAA and detection, powered by an exothermic chemical reaction (Fig. 3b). The reaction, driven by a magnesium-iron alloy and water, is controlled using filter paper to regulate the water flow from the reservoir to the reaction chamber. To maintain optimal temperatures for LAMP-based amplification, paraffin is employed as a PCM to stabilize the temperature in the amplification chambers. The effectiveness of the device was demonstrated by successfully amplifying and detecting Escherichia coli DNA, achieving detection sensitivity down to as few as ten target molecules in the sample [82].

In 2014, Lillis et al. introduced a heater system based on sodium acetate trihydrate (SAT) within a plastic cup, utilizing SAT as the exothermic reaction to generate heat. This system was employed for the first time with RPA as an isothermal amplification method for detecting HIV-1. Sodium acetate trihydrate releases heat through a crystallization process when activated, which occurs when the liquid form of sodium acetate is seeded with powdered SAT, triggering its transition to a solid state [83]. In 2016, Song et al. developed an instrument-free POC molecular diagnostic system for detecting Zika virus in saliva samples. The system consists of two key components: a disposable microfluidic cassette and a chemically heated cup. The microfluidic cassette features four independent multi-functional amplification reactors. The diagnostic process begins with the collection of saliva in a tube, followed by lysis with a buffer solution. The lysed sample is then filtered through an isolation membrane within the cassette, where nucleic acid extraction occurs. Nucleic acids bind to the silica membrane in the reactors, eliminating the need for an elution step and streamlining the workflow. Isothermal amplification, reverse transcriptase-LAMP, takes place directly on the membrane, powered by Mg-Fe pouch format with PCMs. Results are visually detected using a leuco crystal violet dye, which turns violet in the presence of double-stranded DNA. This system can detect less than 5 plaque-forming units of Zika virus per sample in under  40 min [84].

Qiu et al. in 2017, demonstrated that PCR could also be used in POCT devices by employing convective temperature gradients. They developed a rapid molecular detection platform for the H1N1 virus, achieving results within 35 min (Fig. 3c). This method combines a convective polymerase chain reaction with a dipstick assay. A fully disposable, calcium oxide-heated thermal processor heats the capillary tube from the bottom at a fixed temperature, eliminating the need for electrical power. Amplification occurs spontaneously as reagents move through different temperature zones, reducing amplification time by eliminating the need for controlled temperature cycling. Heating the capillary tube to around 95 °C from the bottom initiates amplification, with reagents experiencing different temperature zones associated with DNA denaturation, annealing, and extension stages. Detection of amplification products is done by a rapid one-step nucleic acid dipstick assay, enabling visual detection without instrumentation [85].

In 2020, Udugama et al. developed a tunable and precise miniature lithium heater (Fig. 3f) [86]. The device presented three key features: (1) its adaptable temperature modulation capability overcomes the limitations of conventional chemical heaters, allowing diverse enzyme sets to be used without inhibition; (2) it achieves target temperatures within 1 min, reducing assay times; and (3) its compact size enables portability for complex biological procedures in electricity-free environments. The design of this heater involved utilizing chemical reactions and bubble flow to extract energy from reactive alkali metals, particularly lithium, chosen for its high energy density (~ 222 kJ/mol), ease of handling, and reactivity with water. The shaping and control of lithium’s surface area enabled the creation of a reliable heating by compressing lithium into a channel where it reacts with water to produce heat. These miniature heaters were demonstrated with T4 bacteriophage DNA amplification with RPA at 42 °C, and fluorescence detection of the amplified DNA using a smartphone device. [86]

In 2024, Feng et al. developed a self-heating, RPA-based, integrated lateral flow (LF) strip biosensor consisting of five main components: the LF strip device, connector, RPA reaction chamber, support device, and heating device. Each of these components can function independently or be combined to form a complete system. The LF strip is housed within the LF strip device, while the heating device contains a self-heating mechanism primarily made of calcium oxide, wrapped in non-woven fabric. When exposed to water via pipetting, the self-heating mechanism generates a controlled amount of heat, creating the necessary external temperature environment for the RPA reaction chamber to function effectively [87].

In Table 2, we present an overview of instrument and electricity-free nucleic acid amplification devices that employ water-based exothermic reactions chronically. These devices utilize different types of water-based exothermic reactions involving various compound, such as calcium oxide, magnesium-iron alloy, calcium oxide with sodium carbonate, calcium oxide with aluminum, lithium, and sodium acetate trihydrate.

For the readout of the summarized example devices, various methods have been utilized, including fluorescence, paper-based methods, and colorimetric assays using microfluidic chips. Additionally, among the isothermal amplification techniques integrated into instrument-free, electricity-free nucleic acid amplification devices, two types have been preferred: loop-mediated isothermal amplification and recombinase polymerase amplification. Recombinase polymerase amplification performs isothermal amplification at a low temperature range (39–42 °C), while loop-mediated isothermal amplification operates at a higher temperature range of 60–65 °C [88, 89].

Non-water-Based Self-heating Systems

In addition to water-activated exothermic reactions, other materials have been developed to enable heat generation autonomously without the need for water. These methods include the use of disposable pocket warmers, sunlight, and air-activated hand warmers (Table 3) [90–92]

Table 3.

Non-water-based, self-heating, instrument-free, electricity-free nucleic acid amplification systems

Sample format, pathogen	Detection time, min	Isothermal method	Self-heating element	Type of PCMs	Readout	Limit of detection/sensitivity	Publication year	Refs.
DNA extracted from Bacillus anthracis	90	LAMP	Disposable pocket warmer	Without PCM	Fluorescence and naked eye	1000 copies of Bacillus anthracis pag and capB gene fragments per tube	2010	[90]
Kaposi’s sarcoma-associated herpesvirus DNA	~ 60	LAMP	Sunlight	PureTemp 68	Optical sensors Light-emitting diodes	19 copies per reaction	2018	[91]
RNA extracted from SARS-CoV-2	17, 84, and 90	RT-RPA	Hand warmer pouch	Without PCM	Naked-eye detection by lateral flow strip	94.1% sensitivity, 100% specificity, and 95.8%, accuracy of the microfluidic system	2021	[92]

LAMP loop-mediated isothermal amplification, min minutes, PCM phase change material, Ref reference, RPA recombinase polymerase amplification, SARS-CoV-2 severe acute respiratory syndrome coronavirus 2

In 2010, Hatano et al. employed LAMP for anthrax detection as an example for anti-bioterrorism POCT application. They used a disposable pocket warmer to heat the LAMP reaction, achieving a stable temperature of approximately 60 °C for DNA amplification over about 90 min. Then, amplified DNA could be detected using ambient light [90].

In 2018, Snodgrass et al. introduced the “Tiny Isothermal Nucleic acid quantification sYstem” (TINY), which utilizes a PCM, PureTemp 68, to store heat isothermally and perform LAMP. The authors claimed that its versatility in heat sources, including sunlight and electricity, makes it suitable for both laboratory and field use. They demonstrated this device with the detection of Kaposi’s sarcoma-associated herpesvirus DNA using a fluorescent dye. In this example, users align the lens with the sun by rotating it within a supporting structure, and lens readjustment was necessary one to three times when heating TINY with sunlight, varying based on geographic position and time of year. Afterward, the latent heat from the melted PCM inside TINY maintained the system at an isothermal state for over an hour [91]. Additionally, in 2021, Li et al. employed a hand warmer containing iron powder, salt, and activated carbon, which is activated by exposure to oxygen in the air. They integrated this with a microfluidic chip for detecting viral severe acute respiratory syndrome coronavirus 2 based on CRISPR-RT-RPA with naked-eye detection using a lateral flow strip readout [92].

Discussion and Outlook

Ongoing progress in nucleic acid amplification techniques, particularly INAA methods, presents a promising future for diagnostics that do not require thermocycling equipment. INAAs offer distinct advantages over PCR for POC applications, including the ease of maintaining a single temperature and faster processing times. The integration of self-heating elements into diagnostic tools, inspired by innovations in smart food packaging, marks a significant leap forward in technological advancement, as discussed through the examples in this review. Expanding these self-heating technologies beyond their traditional use into the realm of molecular diagnostics could revolutionize personalized healthcare.

The integration of exothermic reactions with PCMs in portable diagnostic tools, such as NINA devices, holds significant potential to improve diagnostic accessibility, particularly in resource-limited settings. By reducing turnaround times and enabling efficient, portable diagnostics, these innovations could lead to the development of fully integrated ‘sample-to-result’ platforms that can be operated by minimally trained personnel, thereby greatly enhancing healthcare efficiency and accessibility. Furthermore, the incorporation of self-heating elements in diagnostic systems could facilitate the development of electricity-independent platforms for on-site detection and at home use, offering a profound impact on global health security, particularly in response to the increasing threat of emerging infectious diseases. Future directions involve the commercialization of these electricity-free, nucleic acid-based devices, similar to the self-heating food and beverage products available to consumers. This approach not only facilitates the rapid and accurate identification of active infections but also holds great promise for the development of multiplexed assays, allowing for the simultaneous detection of multiple disease types.

However, commercializing these self-heating diagnostic tools poses significant challenges. Regulatory hurdles remain a critical concern, as these devices must meet stringent safety and efficacy standards before gaining approval for medical use. While self-heating components in food packaging have already been approved for commercial use, transitioning these technologies into medical applications will require additional regulatory scrutiny. Moreover, cost considerations will be a factor in determining the widespread adoption of these devices, as manufacturers must balance affordability with the sophisticated technology involved. Market acceptance will depend on demonstrating the reliability, safety, and value of these diagnostics to both healthcare professionals and consumers.

The SWOT analysis of self-heating elements in the smart food technology market highlights several key factors. The strengths of this technology include its significant convenience and portability, making it well suited for on-the-go lifestyles, outdoor activities, and emergency situations. Its ability to deliver consistent sufficient heating without the need for external appliances distinguishes it in a competitive market. However, high development costs and stringent regulatory requirements may limit growth and adoption. Opportunities for expansion are driven by ongoing technological advancements and the increasing demand for convenience. Progress in chemical and material sciences presents possibilities for enhancing safety, sustainability, and cost efficiency, positioning this technology for future growth. Nonetheless, the market also faces threats, such as intense competition from established players and a reliance on continued technological innovation. Successfully navigating these challenges will require balancing innovation, safety, sustainability, and cost effectiveness to capitalize on opportunities and address the inherent risks.

Nevertheless, the applications of these electricity-free and instrument-free diagnostic devices are endless and not limited only to regular pathogen monitoring in clinics, hospitals, pharmacies, and even homes. During outbreaks, these devices could play a crucial role in quickly and accurately detecting pathogens, helping to prevent the spread of disease. In industries such as food production, this approach could be used to monitor pathogens such as Salmonella, Legionella pneumophila, Mycobacterium avium, Pseudomonas aeruginosa, and many others in drinking water, addressing severe health risks. Additionally, in environmental monitoring, these tools could facilitate on-site pathogen detection without the need for sample transport, enabling immediate action. As these technologies evolve, their potential extends even further, including the detection of bioterrorism threats and ensuring public safety. By addressing the challenges of cost, regulation, and market acceptance, self-heating diagnostic tools have the opportunity to revolutionize POC diagnostics, particularly in resource-limited and remote settings. With continued innovation and exploration of new applications, these devices can meet the ASSURED criteria set by the World Health Organization, ensuring that diagnostics are affordable, sensitive, specific, user-friendly, rapid, robust, equipment-free, and deliverable to end users.

Declarations

Funding

Open access funding was enabled and organized by Projekt DEAL.

Conflicts of Interest

Mojdeh Hamidizadeh, Renata F. Martins, and Frank F. Bier have no conflicts of interest that are directly relevant to the content of this article.

Ethics Approval

Not applicable.

Consent to Participate

Not applicable.

Consent for Publication

Not applicable.

Availability of Data and Material

Data sharing is not applicable to this article as no datasets were generated or analyzed during the current study. All data generated or analyzed during this study are included in this article.

Code Availability

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Authors’ Contributions

MH: idea, writing, review, and editing; RFM and FFB: review and editing.