COVID-19: Researchers have developed a highly accurate test that can analyze hundreds of samples at the same time


Antibody testing can be a powerful tool for tracking the spread of SARS-CoV2 infections, the virus responsible for the COVID-19 pandemic. A group of scientists from EPFL, UNIGE and HUG have now developed a reliable and cheap antibody test that can analyze more than 1,000 samples at once and requires a small drop of blood, such as that from a finger prick.

After people get infected with the SARS-CoV-2 virus that causes COVID-19, they start to produce immune molecules called antibodies. COVID-19 antibody tests pick up on the presence of antibodies against SARS-CoV-2 in the blood. Because antibodies can take several days to weeks to develop, antibody tests can’t detect active infections, but they can help to find out what proportion of communities have been infected with the virus in the past.

This knowledge is useful for epidemiological investigations, and informing public-health policies. Antibody tests are also a powerful tool to evaluate COVID-19 vaccine efficacy in clinical trials, when scientists look at the rise in antibodies after volunteers get a jab.

However, antibody tests rely on rather expensive reagents and typically require larger quantities of blood taken with a venous blood draw, which can only be performed by trained healthcare personnel. What’s more, some of the tests on the market are too inaccurate to deliver reliable results. Now, researchers from EPFL, UNIGE and HUG have developed a highly accurate test that can analyze hundreds of samples at the same time, using minute quantities of reagents and single drops of blood.

“The coolest thing about our approach is that you can do a lot of tests at once with minimal reagents, and you could even have people collect their own blood samples at home,” says study first author Zoe Swank, a former Ph.D. student in the EPFL’s Laboratory of Biological Network Characterization led by Sebastian Maerkl.

In early 2020, Swank and Maerkl teamed up with Benjamin Meyer, a virologist at UNIGE Faculty of Medicine and scientific collaborator at HUG Division of Laboratory Medicine, and with Isabella Eckerle, a professor at UNIGE Faculty of Medicine and Medical Coordinator of the UNIGE-HUG Centre for Emerging Viral Diseases, and set out to repurpose a diagnostics platform that had been previously developed in Maerkl’s lab, so that it could be used to perform SARS-CoV-2 antibody tests.

The platform, which can analyze up to 1,024 samples at once, consists of a complex network of tiny tubes carved into a plastic chip that is about the size of a USB stick. To perform the assay, the researchers feed individual blood samples and test reagents through the channels of this ‘microfluidic’ chip. If antibodies against SARS-CoV-2 are present in a blood sample, a molecule generates a signal that can be detected as a fluorescent glow under a microscope.

When the team tested blood samples from 155 individuals infected with SARS-CoV-2, the assay detected antibodies against the virus in 98% of cases. The assay is also extremely specific: it never detected antibodies against the virus in samples from people who had not been infected with SARS-CoV-2.

COVID-19 test detects antibodies in hundreds of tiny blood samples
A MITOMI microfluidic device © Sebastian Maerkl / 2021 EPFL

Because the microfluidic device is very small, the amounts of blood and reagents used are a fraction of those required for standard COVID-19 antibody tests. And running hundreds of assays on a single platform means that a person can perform more assays in less time, with potential cost savings on human labor, Maerkl says. “If you do a back-of-the-envelope calculation and take everything into consideration, including salary costs and the cost of reagents, it is about 0.5 Swiss francs per assay,” he says. “It’s almost negligible.”

To eliminate the need for collecting blood from people’s veins, Swank and her colleagues assessed whether they could use blood samples obtained from a finger prick—a simple procedure in which a finger is pierced with a tiny needle to obtain a small quantity of blood. The researchers tested three commercially available devices to perform finger-prick blood tests, including glucose test strips used by people with diabetes to measure their sugar blood levels.

The microfluidics-based antibody test could be successfully run on blood samples collected with all three methods, even when the blood was left to dry and stored for about one week at room temperature, or when samples were shipped by regular mail from Geneva to Lausanne. The study was published in PNAS.

“The approach of collecting blood in a decentralized way by a simple finger prick that can be even done at home, and a sophisticated laboratory-based assay with high diagnostic accuracy makes this test very attractive for large-scale epidemiological studies, explains Isabella Eckerle.

It could even be used for remote geographic regions that lack sufficient laboratory capacity, for example to conduct seroprevalence studies in Sub-Saharan Africa.” She adds that “the small amount of blood and the collection by a finger prick, which is quick and almost painless, also makes this method very attractive for the use in children and offers a unique opportunity to assess seroprevalence rates in daycare centers or kindergartens.”

Maerkl and his collaborators are now using the test to determine the prevalence of antibodies against SARS-CoV-2 among kindergarteners in Geneva, in collaboration with Silvia Stringhini and Idris Guessous of the Population Epidemiology Unit at HUG. In the future, Maerkl says, this technology could make it possible for people to buy a blood sampling kit at a pharmacy or a supermarket, collect their own blood with a simple finger prick, and mail it to a central laboratory that analyzes the blood sample and returns the test results via email or a smart-phone app.

There’s no obvious limit on how many molecular assays can be done using the microfluidic diagnostics platform, Maerkl adds. “We’re interested in expanding this platform to other types of assays that would detect other biomarkers that people might want to measure—for example, blood ferritin levels in people with anemia,” he says.

The emergence of a new coronavirus at the end of 2019, termed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), led to an unprecedented global public health crisis (1). Over a year later, it is estimated that SARS-CoV-2 infected over 100 million people worldwide, and 2 million people died of COVID-19, caused by SARS-CoV-2 (2).

As SARS-CoV-2 causes mainly mild disease or infection presents without symptoms, many cases are not captured by direct testing in the acute phase of disease (3). However, to estimate infection fatality rate and guide public health decisions, it is of utmost importance to establish the true spread or prevalence of the virus by identifying how many people have been exposed (4, 5).

Detection of anti–SARS-CoV-2 antibodies using highly sensitive and specific assays can help answer these questions. Several seroprevalence studies have already been conducted, demonstrating rather low seroprevalence rates even in areas that were severely affected (6⇓⇓–9). Such data indicate that herd immunity through natural infection is far from being reached. However, these studies are merely snapshots of an evolving situation, both in time and space.

Therefore, there is a sustained need for seroprevalence studies to be continuously conducted in order to monitor virus spread and to keep policy makers informed. In addition, tens to hundreds of thousands of blood samples will need to be tested for each SARS-CoV-2 vaccine phase 3 clinical trial, and large numbers of samples will need to be tested to monitor immune responses after a vaccine has been approved and rolled out.

Such studies are cumbersome and expensive to perform, as they require large numbers of serum samples obtained by venipuncture and analyzed by highly sensitive and specific immunological assays to classify samples as seropositive or seronegative.

Several assays, such as enzyme-linked immunosorbent assays (ELISA) or chemiluminescent immunoassays (CLIA), are commercially available, but mainly rely on serum drawn by venipuncture. These tests are also rather expensive, with reagent costs on the order of 3 USD to 10 USD per test. Alternatively, in-house ELISAs are difficult to standardize and require high amounts of recombinant antigen, usually around 100 ng per sample (5).

Other recently developed methods such as miniaturized high-throughput ELISAs that use low microliter volumes suffer from lower sensitivity (10), and ultrasensitive assays based on digital ELISA have low sample throughput of 68 tests per hour (11). The comparatively high cost of these assays and the reliance on serum samples taken by venipuncture are considerable hurdles to performing large-scale studies under normal circumstances, but especially so during a pandemic, when sample collection can put clinical staff and study participants at risk.

Lateral flow assays (LFAs) can be performed at the point of care or at home, requiring only a “drop” of whole blood, but the sensitivity and specificity of these assays is often low (12⇓–14), and LFAs are relatively expensive, at ∼22 USD per test. Furthermore, LFAs provide test results, but no blood samples are being collected which could be used for follow-up analyses.

There is therefore a need for new technologies to supersede existing methods such as ELISA, CLIA, and LFAs. Novel technologies should be capable of high throughput, low reagent consumption, and low cost per test; achieve high sensitivity and specificity; and be compatible with ultralow-volume whole blood samples in the low or even submicroliter range that can be obtained via a simple finger prick.

Biomarker detection using dried whole blood on filter paper or other devices would have tremendous advantages, as the sample can be collected by untrained individuals at home. The samples could then be conveniently shipped by regular mail at ambient temperature to a central laboratory for analysis, and test results could be returned electronically via a mobile app or email.

In this study, we developed and validated a nanoimmunoassay (NIA) that analyzes 1,024 samples in parallel on a single microfluidic device the size of a USB stick. NIA reagent consumption and corresponding costs are roughly 1,000 times lower than a standard ELISA. NIA achieved a specificity of 100% and a sensitivity of 98%, based on the analysis of 134 prepandemic negative sera and 155 positive sera from RT-PCR–confirmed positive individuals.

NIA performed well for samples obtained more than 20 d post onset of symptoms, and performed equally well for samples obtained less than 20 d past onset of symptoms. We go on to demonstrate that NIA can be used to detect anti–SARS-CoV-2 antibodies in ultralow-volume dried whole blood samples, eliminating the need for venipuncture blood collection. We tested two commercial blood collection devices: Neoteryx’s Mitra® and DBS System SA’s HemaXisTM DB10, and show that it is possible to repurpose low-cost and widely available blood glucose test strips for sample collection and shipment. Samples could be stored up to 6 d at room temperature with minimal sample degradation. All three methods combined with NIA identified more positive samples than a standard ELISA performed on serum samples collected from the same individuals.

High-throughput microfluidic NIA for anti–SARS-CoV-2 antibody detection. (A) The two-layer microfluidic chip design consists of 1,024 unit cells. Each unit cell, in turn, contains two sections: an immunoassay chamber (top) and a spotting chamber (bottom). Control valves include the button (1), sandwich (2), and neck (3) valves. (B) A schematic of the experimental process, starting with the spotting of patient samples, followed by chip alignment and bonding, biotin-BSA and neutrAvidin surface patterning, and, lastly, the immunoassay for detection of anti–SARS-CoV-2 antibodies. Surface patterning and immunoassay are shown for a single unit. During the experiment, all unit cells are processed in parallel. (C) Schematic of the on-chip sandwich immunoassay. (D) Fluorescence images of anti-human IgG-PE signal for a given concentration of anti-spike antibodies present in human serum. (E) Quantification of the anti-human IgG-PE signal for a range of anti-spike concentrations spotted. The dashed horizontal line indicates the LOD.

reference link:

More information: Zoe Swank et al. A high-throughput microfluidic nanoimmunoassay for detecting anti–SARS-CoV-2 antibodies in serum or ultralow-volume blood samples, Proceedings of the National Academy of Sciences (2021). DOI: 10.1073/pnas.2025289118


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