The diagnostic tool creates stronger, longer bioluminescence signals for virus detection

Mass General Brigham researchers are sheding a powerful new light into the viral darkness with the development of a sensor based on the luminescent CAscade (LUCAS), a fast, portable, highly sensitive diagnostic tool for processing complex biological samples.

Compared to its diagnostic predecessors, LUCAS creates 500 times stronger and eight times longer bioluminescent signals, overcoming the long-standing problems faced by on-site diagnostics. Their study was published in the journal Nature Biomedical Engineering.

"The development of effective diagnostics is incredibly difficult, especially if we think about the particle size of infectious diseases and complex biological fluids in which we try to identify them. Finding an HIV particle in a human blood sample is like finding an ice cube in a jelly-filled Olympic pool with blindfolded eyes," said senior author Hadi Shafi, Ph.D., lecturer at the Faculty of Medical Engineering and the Department of Renal Medicine at Brigham and Women's Hospital.

"Thanks to its new enzyme cascade approach, LUCAS marks a significant leap forward for probing viruses in these complex biological samples."

On-site first aid diagnostics have become important tools in many households, as people measure blood sugar levels, take pregnancy tests and even conduct their own tests for COVID-19. These diagnostic procedures, which allow people to avoid tedious, expensive laboratory tests, are important for the detection, treatment and monitoring of diseases.

However, the current diagnosis may fail, with such shortcomings as inaccuracy and poor sensitivity. Bioluminescence can alleviate common defects experienced by other methods, such as background noise, false positive results, photobleaching and phototoxicity.

Bioluminescence uses the same natural enzyme that causes fireflies to glow to illuminate biological samples for visualization. The enzyme, luciferase, is added to the sample to search and label viral particles. Then luciferin molecules are introduced into this sample, causing a luciferase reaction that creates a burst of light. But this reaction produces a light signal that is both weak and short-lived.

Shafi and his team have developed a unique cascade of enzyme signals to enhance and extend bioluminescence signals. They introduce another enzyme called beta-galactosidase into the equation, which sticks to luciferin and releases it continuously, instead of allowing luciferin to float freely in the sample for one-time reactions.

This additional step means more luciferin, more luciferase reactions and more bioluminescence. In fact, this step allowed LUCAS to be 515 times more bioluminescent than systems without LUCAS, and LUCAS signals remained strong at 96% after an hour.

To assess the effectiveness of LUCAS, the team used 177 viral samples of patients and 130 viral serum samples infected with SARS-CoV-2, HIV, HBV or HCV. Samples of patients with SARS-CoV-2 were taken with a nasopharyngeal swab, while samples of HIV, HBV and HCV were collected using blood sampling. LUCAS provided diagnostic answers within 23 minutes and with an average accuracy for all pathogens of more than 94%.

Researchers have designed LUCAS both portable and easy to use so that it can be an option for high and low resource environments. As a next step, the team will check the effectiveness of LUCAS in other biological fluids and whether the method can identify more than one pathogen at the same time.

Shafi also notes that the identification of biomarkers for many diseases, including Alzheimer's disease, is a rapidly developing space, so the presence of a tool such as LUCAS, ready to work as new biomarkers emerge, may have an impact in the coming years.

"We always want to detect infection and diseases as early as possible, as it can make a big difference when it comes to care and long-term results," said the first author Sungwan Kim, Ph.D., a doctoral student at the Shafi Laboratory in Brigham.

"With our focus on developing sensitive, accurate and accessible diagnostic tools, we want to make early detection easier than ever and take personalized care into a new era."