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Lab-On-A-Chip Device Taking On One Of The Biggest Diseases Of Poverty

By Deborah Borfitz 

July 18, 2023 | Capturing tuberculosis (TB) bacilli in a sputum sample can be like finding a needle in a haystack, unless the bacterial load is high. That’s when most cases are currently detected in low- and middle-income countries (LMICs) around the world. The “missing millions” go on to become part of the seemingly unmovable World Health Organization (WHO) statistic of 1.4 million deaths annually, many of whom are children (250,000), according to Philip D. Butcher, Ph.D., professor of molecular medical microbiology at St. George’s University of London. 

Butcher calls the situation immoral and unjust, as well as largely invisible to people in the prosperous Western world where access to healthcare is a given and TB is largely controlled. “Because it’s not in the mainstream [and] politicians’ families aren’t dying from TB, nobody cares about it. It’s a Third World problem,” he says.  

The current scenario in LMICs are patients presenting in the hospital with symptoms of active TB— including fever, night sweats, and persistent coughing—who are then evaluated by X-ray and confirmatory lab tests. “A huge amount of TB is continually transmitted in high-incidence countries because people aren’t diagnosed quick enough or early enough,” says Butcher. By the time patients seek care they may have already infected other people for many months, he notes. “Having a test that you can take out of the hospital environment and into a community-based setting would allow these early diagnoses to occur.” 

The enabling technology, Butcher says, is unlike anything a hospital lab might use—specifically, a chip-based diagnostic test called CAPTURE-XT. The test is based on dielectrophoresis to selectively isolate the causative pathogen Mycobacterium tuberculosis (Mtb) for visual detection.  

Ensuring equitable access to diagnostics for this killer disease, thereby controlling its unintentional spread, is a scientific endeavor for Butcher. For much of the past eight years, he has been working closely on the development of this cheap, rapid, accurate, and portable TB diagnostic test with QuantuMDx (Newcastle upon Tyne, England).  

Particle Capture

Currently, half of all TB testing is sputum smear microscopy, and that paradigm is overdue for disruption, says Jonathan O’Halloran, Ph.D., founder and CEO of QuantuMDx. It is entirely possible that a highly sensitive dielectrophoretic chip could one day replace the slides and stains and required clinical expertise associated with smear microscopy for roughly the same price.  

As was recently demonstrated by the research team in a study that published in The Journal of Molecular Diagnostics (DOI: 10.1016/j.jmoldx.2023.04.005), the prototype device can process sputum from suspected TB patients, capture Mtb bacilli, and provide a purified sample for molecular confirmation by quantitative PCR and ultimately for genotypic drug-susceptibility analysis. This suggests the technology could serve as a front-end sample preparation tool for enhanced molecular detection not reliant on a central laboratory. 

But the longer-term goal is a point-of-care test that would use basic optics to detect Mtb as it is being collected by a PCR assay using electric probes. In an upcoming paper, QuantuMDx will be showing how CAPTURE-XT can be used on its own to culture TB cells and observe them dividing, and then being killed with identified antibiotics at different concentrations, O’Halloran says. “We’ll be looking [on the fly] at the right dosage of a drug for the mycobacteria” based on changes to an individual’s dielectrophoretic profile.  

Cells of different shapes, sizes, and permittivity (ability to store electrical energy) can be captured in a dielectrophoretic field of a certain frequency and magnitude, explains O’Halloran. Importantly, TB bacilli can be captured in this way, but the same technology could be used to find circulating tumor cells or bloodstream infection bacterium and fungi. The possibilities, most notably for TB and sepsis, are being investigated in parallel, he adds. 

Dielectrophoresis has previously been deployed in the agricultural industry for visualizing and detecting disease-causing organisms, and electrophoresis—a related electrokinetic particle manipulation technique that separates only charged particles—has on occasion been used for sample preparation purposes in human-based disease diagnoses, says Butcher. But this may be the first-ever application of dielectrophoresis to clinical microbiology in a chip-based system.  

Impressive Results

Dielectrophoresis is a technique that uses the principles of polarization and the motion of bioparticles in applied electric fields. It can be tuned to selectively attract or repel specific particles or cells based on their dielectric properties, a measure of their potential for charge movement in response to an external electric field, says O’Halloran. Here, Mtb bacteria is being captured and concentrated, while the other sputum contents are washed away. 

For the latest published study, the dielectrophoretic chip was first optimized using a panel of 50 characterized sputum samples. Its performance was then assessed by a blinded screening of 100 characterized and bio-banked sputum samples provided by the Foundation for Innovative New Diagnostics. 

Concordance with culture diagnosis was 100% for smear negative samples and 87% for smear positive samples. Of the smear positive samples, the high burden sample concordance was 100%.  

Those results are a bit misleading, since the gold standard against which the chip system was compared produces false negatives, says Butcher. Sputum smear microscopy requires as many as 10,000 Mtb bacilli per milliliter to detect TB infection. “The numbers under-estimate how good we are.”  

The initial dilemma, and focus of preliminary experiments, was how to capture only Mtb bacilli in sputum—a complex specimen type full of other bacteria, proteins, and cells from the immune system, he continues. The latest paper includes a video showing the flow of sputum through the microfluidic chamber passing over the primary electrode bed, and the gradual trapping of the smaller bacilli particles onto the edges of the dielectrophoresis electrodes.  

Road To Market

The current reality is that two to three million people each year fail to be diagnosed with TB, says Butcher, primarily in the 13 “high priority” countries identified by the WHO, Stop TB Partnership, and the United Nations. These are located mainly in Sub-Saharan Africa, Southeast Asia, and parts of Eastern Europe where health resources are poor as is access to healthcare. 

It’s unlikely that CAPTURE-XT will compete with lab-based diagnostic systems that are commonplace and free or affordable in wealthier countries of the world, he adds. The target audience are those nations with high levels of TB who have been missing out on standard diagnostics for various reasons, including access issues as well as the stigma associated with being diagnosed with TB in a hospital. 

In LMICs, QuantuMDx will take CAPTURE-XT through the WHO prequalification program for in vitro diagnostics as well as the licensing programs of the individual countries, O’Halloran says. Validation studies will also be needed at the hospital and laboratory level. “It’s a long pathway, it is not optimized, and it’s expensive ... [but] at the end of it you get a high-quality test that has been rigorously tested and by the time it hits the market a lot of the implementation work has been performed.” 

The prequalification studies will take another three or four years to complete, says O’Halloran, and will be done in-country. Some initial studies already conducted in South Africa have shown “very good results,” he adds.  

Presumably, some of the large advocacy organizations that are desperately trying to reduce the worldwide TB burden (including the Bill & Melinda Gates Foundation and the Stop TB Partnership’s TB Reach program) would help support the initial rollout of CAPTURE-XT to these high-priority countries. “There are still an incredible 10 million new cases in the world every year... and those numbers haven’t really changed much over the last 15 years,” says Butcher. “Disruptive technologies are what it is going to take to do something about it.”