Simple, inexpensive and sensible | Nature Biomedical Engineering
Designing viral diagnostic tests for point-of-care use involves many trade-offs. Yet ease of use, low cost and accuracy should not be compromised.
An ideal point-of-care test to check for infection (or contagiousness) should be accurate, affordable, and accessible. Accuracy implies high sensitivity and specificity for the infectious agents being tested, and accessibility implies that the test is quick, easy to use, and available where it is needed most. These are the three main attributes of the ASSURED criteria, which were established by the World Health Organization’s Special Program for Research and Training in Tropical Diseases in 2003 to help expand access to point-of-care diagnostics. service. ASSURED means affordable, sensitive, specific, user-friendly, fast and robust, without equipment and deliverable to end users. Two additional timely criteria — real-time connectivity and ease of sample collection — and the acronym REASSURED were proposed in 2019 (ref. 1).
Credit: Ting Zhang and Chengyong Wu, Sichuan University
The realization of an ideal is almost always accompanied by substantial compromises. Accurate tests, especially nucleic acid tests using the gold standard polymerase chain reaction (PCR) technology for nucleic acid amplification, involve expensive laboratory instruments, are not immediately or widely available, and are carried out by qualified experts. Accessible and affordable tests, such as over-the-counter tests for antigen detection, may be relatively inexpensive and easy to use, but are not as sensitive and may not be robust. And rapid antibody tests may be insufficiently specific.
The relevant trade-offs and how they inform the design and implementation of a point-of-care test depend on the intended use of the test and the conditions under which it is used. For example, for respiratory infections, an insufficiently sensitive but inexpensive test may be sufficient to quickly verify contagiousness (but not for active infection), and may be supplemented by a more accurate and expensive confirmatory test. A test that can test multiple viruses or virus strains simultaneously may be more complex to use and less affordable, but suitable in the context of a hospital or doctor’s visit. And an easy-to-use, accurate test that requires equipment or reagents that must be stored in refrigerators may not be accessible enough to use in low-resource settings.
Yet, regardless of usage parameters, designing diagnostic tests for truly widespread use at the point-of-care should not compromise usability, affordability, or accuracy. Tests that are slower to provide results but require only basic hardware equipment are more likely to be useful and easier to adopt in health and public health settings, even if the tests are not as robust (as long as they implement internal controls), as rapid, robust tests that are not as accurate or affordable or require user training. Indeed, the usefulness of home lateral flow antigen testing in determining when someone infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) should self-isolate has been overlooked for too long.2. Four articles included in this issue of Nature Biomedical Engineering provide solutions to some of the design and engineering challenges that prevent adapting and simplifying tests to make them usable at the point of service.
In an article, Cameron Myhrvold and his collaborators tell how they adapted their3 Cas13-based test for detection of SARS-CoV-2 to facilitate test use and deployment. The test does not require the extraction of RNA from a nasal sample, involves freeze-dried reagents and rapid inactivation of ribonucleases at room temperature, can be performed using body heat to catalyze reactions, and the result can be read on a strip of paper; therefore, heating or cold storage equipment and optical lenses and detectors are not strictly necessary, making the test relatively inexpensive. In addition, the test is significantly more accurate (100% specific and 90.5% sensitive when compared to reference PCR) than equally rapid commercial antigen tests, and can be easily adapted for the discrimination of viral variants of concern. However, despite these capabilities and optimizations, the test does not meet the ASSURED criteria. As explained in an accompanying News and Views article by Ahmed Ghouneimy and Magdy Mahfouz, the test involves manual handling of liquids and five simple steps, and would therefore require additional design optimizations to facilitate performance by an untrained person. .
In another article, Patrick Hsu, David Savage, Jennifer Doudna and colleagues also report a sensitive and rapid Cas13-based test designed for point-of-care use that can detect multiple SARS-CoV-2 variants in saliva samples. and it doesn’t work. t requires extracting RNA from the sample (but requires it to be denatured at high temperature). The test incorporates an internal control and uses a microfluidic cartridge (single-use, gravity-driven) that automatically performs liquid handling operations. However, processing the sample before introducing it into the device involves two manual steps. Additionally, the need for custom equipment (the cartridge and its housing device incorporating a compact fluorescence reader) is a barrier to the affordability and scalability of the test.
A third paper, written by Jinghong Li, Weimin Li, Ruijie Deng and co-authors, reports a test that dispenses with nucleic acid amplification and colorimetrically detects several specific RNAs simultaneously (as shown by the authors for various circulating strains of SARS-CoV-2) by taking advantage of strand displacement reactions (via a DNA exchange probe) and enzymatic amplification of the detection event (via the release of Ag(I) ions , which hydrolyse urea and change the pH and color of the solution). The test is quick (the result is obtained in 30 min) and sensitive, and the reaction steps are implemented on inexpensive origami paper (pictured) and performed by folding it. As Kaiyue Wu and Alexander Green pointed out in an accompanying News and Views article, the test combines the specificity of nucleic acid tests with test times and costs that approximate those of lateral flow tests. . However, the test may not be sensitive enough to detect infections early, and the actual performance of the paper folding steps may not be robust enough for widespread use.
The ASSURED criteria do not directly consider the point-of-care benefits of testing that incorporates multiple tests. Donald Ingber, James Collins and colleagues argue, in a paper also included in this issue, that a multiplexed electrochemical test on a chip can help quantify rates of SARS-CoV-2 reinfections and declining levels of protection provided by vaccination. The article describes the integration of simultaneous nucleic acid and antibody testing for SARS-CoV-2 RNA in saliva and for immunoglobulins in blood plasma. The test runs within two hours and has single-molecule sensitivity, and the device extracts, concentrates and amplifies SARS-CoV-2 RNA from untreated saliva. To make it affordable and more accessible, the lab-on-a-chip should be reusable and robust, and fully integrate electronics, pumping and readout systems.
Tests that meet all or most of the ASSURED criteria exist for the detection of antibodies or antigenic biomarkers of HIV, malaria, syphilis, tuberculosis and a few other infectious pathogens1. Inexpensive and accurate home-use tests that detect infection with endemic respiratory viruses should follow. Yet tests that warn of virus exposure2 in real time and without user intervention, and therefore before the actual infection, would be really reassuring.
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Nat. Biomedical. Eng 6, 923–924 (2022). https://doi.org/10.1038/s41551-022-00935-w