Devices made with cheap strips of paper have outperformed two other testing methods in detecting malaria infection in asymptomatic people in Ghana - a diagnostic advance that could accelerate efforts to eliminate the disease, researchers say.
Deceptively simple in appearance, the devices facilitate chemical reactions between a drop of blood and molecules embedded into paper layers and rely on sophisticated, but portable, instrumentation to make the diagnosis: a mass spectrometry measurement of the final product - in positive cases, a malaria-specific antigen that triggers the immune system.
"Typically you would take the sample to the lab, but now we are taking the lab to the sample - I'm taking it to Africa, one of the remotest parts of the world, and doing the analysis right there," said Abraham Badu-Tawiah, lead author of the field study report and professor of chemistry and biochemistry at The Ohio State University.
"The question was, can we have a sensitive tool that can be delivered to people no matter where they are. Statistical analysis showed that our method is 90% accurate, comparable to a PCR test. It's very good and we can deliver these results to people who need it the most."
The research was published recently in Analytical Chemistry.
Malaria is caused by the bite of mosquitoes that spread infectious parasites. The World Health Organization estimates that in 2022, 249 million people globally had malaria, and about 608,000 died of the disease. A preventive vaccine is now available to children in Ghana, where over a quarter of the population was infected in 2011 compared to 8.6% by 2022.
Badu-Tawiah first reported on this invention in 2016, describing a device for at-home or remote-location testing using lightweight structures that could keep biological samples stable for months at a time.
Though the technology is already being refined for the detection of other diseases, malaria has been Badu-Tawiah's chief concern - especially as increased uptake of the vaccine lowers natural immunity among the population, creating the need for widespread surveillance for potential infections in sub-Saharan Africa.
Since 2016, Badu-Tawiah's lab has created a 3D automation process of storing antibodies and ions in the device and added a multipronged molecule to amplify the compound signal for detection by mass spectrometry, but the device fabrication process is still manual. Sheets of paper composing the device's layers - coated with waxy sections that keep blood from seeping through - are printed individually and pressed together with double-sided tape. Twenty-five devices fit onto the 8x12-inch sheets.
Once applied, the blood is separated into four chambers - two acting as positive and negative controls - and induces chemical reactions as it passes through the layers. The chemists designed ionic probes to tag antibodies that extract the antigen from the blood and place it permanently onto the paper within about 10 minutes. Following a buffer wash, the strips are peeled apart and waved in front of a handheld mass spectrometer.
"The spectrometer measures the mass of the compound of interest. The molecular weight tells us if we see a specific mass, that means the malaria antigen is in your blood. That's a yes. If it's not there, that's a no," Badu-Tawiah said.
Results are available in about 30 minutes, but used devices can also be stored indefinitely without refrigeration for later analysis. The high stability means that after the washing phase, the devices can be transferred in ordinary envelops - a capability connecting people with asymptomatic infection in the remotest regions of Africa to resource-rich centers anywhere else in the world, without traditional cold-chain restrictions.
Over five weeks in 2022 in Ghana, Badu-Tawiah tested the device's effectiveness in 266 asymptomatic volunteers and compared its results to three other common testing methods in current use for malaria diagnosis: microscopic examination of blood cells, commercially available rapid diagnostic tests and PCR (polymerase chain reaction).
A key factor in testing people without symptoms, Badu-Tawiah noted, is that if they are infected, the density of parasites in their blood is likely low - meaning a highly sensitive test is needed to detect their presence.
The comparison showed that microscopy, the gold standard in African hospitals, had the least accurate results, indicating only 24 positive cases, and rapid diagnostic tests identified 63 infections. PCR identified 142 positive cases, and the paper-based devices identified 184 positives.
"Microscopy works well when the person is sick and in the hospital. Here, we were in communities, where only 24 were shown as positive with microscopy - this test is telling us the majority are negative. That's a big problem," Badu-Tawiah said. "But when using a more sophisticated method like PCR, almost 50% of the people are sick, and yet microscopy can't tell us that. And in people with very low parasite density, rapid diagnostic tests failed miserably - they can only detect higher parasite density."
Calculation of each method's sensitivity - the number of true positives divided by true positives plus false negatives - showed that the paper-based devices reached 96.5% sensitivity, compared to 17% for microscopy and 43% for rapid diagnostic tests.
Forty-seven out of 266 samples gave a false-positive result - and all were confirmed by microscopy to be negative. PCR, which is considered to be the most accurate test, also diagnosed these people as negative.
Badu-Tawiah said the false positives could have been caused by differing viscosity of the blood samples, leading to redistribution of blood channels during the washing phase. The team modified the device to account for that possibility.
Badu-Tawiah has begun conversations with the Ghana government about implementation of a testing program.
"We told people this was possible in 2016, and we've actually gone to the field and tested it. It's very promising," he said. "Technology will go hand-in-hand with vaccination, and you need a sensitive tool that is deliverable."
He is also partnering with clinicians at Ohio State on adapting the devices to detect risks for, among other conditions, colorectal cancer and acute pancreatitis.
"I have the hammer now and I could hit different nails," he said. "All we have to do is change the antibody to make it applicable to other diseases."
This work was supported by the National Institute of Allergy and Infectious Diseases. Co-authors include Ayesha Seth, Suji Lee, Girish Muralikrishnan, Edgar Garcia and James Odei of Ohio State, and Abdul-Hakim Mutala and Kingsley Badu of Kwame Nkrumah University of Science and Technology in Ghana.
