Like the COVID-19 pandemic continues to spread around the world, and testing is still an important strategy to detect and contain the virus. Maga Alafeef, a graduate student in bio-engineering, co-developed a rapid, ultra-sensitive test using a paper-based electrochemical sensor that can detect the presence of the virus in less than five minutes. The team led by Professor Dipanjan Pan presented their findings in ACS Nano.
“Currently, we are experiencing a once-in-a-lifetime life-changing event,” Alafeef said. ‘We respond to this global need from a holistic approach by developing multidisciplinary tools for early detection, diagnosis and treatment for EARS-CoV-2. ”
There are two broad categories of COVID-19 tests on the market. The first category uses reverse transcriptase real-time polymerase chain reaction (RT-PCR) and nucleic acid hybridization strategies to identify viral RNA. Current FDA-approved diagnostic tests use this technique. Some disadvantages include the amount of time it takes to complete the test, the need for specialized staff and the availability of equipment and reagents. The second category of tests focuses on the detection of antibodies. However, it can take a delay of several days to a few weeks after someone has been exposed to the virus to produce detectable antibodies.
In recent years, researchers have been successful in creating careful biosensors using 2D nanomaterials such as graphene to detect diseases. The main benefits of graphene-based biosensors are their sensitivity, low production cost and fast detection. ‘The discovery of graphene opened a new era of sensor development due to its properties. Graphene exhibits unique mechanical and electrochemical properties that make it ideal for the development of sensitive electrochemical sensors, ”said Alafeef. The team created a graphene-based electrochemical biosensor with an electrical readout setup to selectively detect the presence of SARS-CoV-2 genetic material.
There are two components in this biosensor: a platform to measure an electrical readout and probes to detect the presence of viral RNA. To create the platform, researchers first coated filter paper with a layer of graphene nanoplates to create a conductive film. Then they placed a gold electrode with a predetermined design on top of the graphene as a contact block for electrical reading. Both gold and graphene have high sensitivity and conductivity, making this platform ultra-sensitive to detect changes in electrical signals.
Current RNA-based COVID-19 tests for the presence of the N gene (nucleocapsid phosphoprotein) on the SARS-CoV-2 virus. In this study, the team designed antisense oligonucleotide (ASOs) to target two regions of the N gene. The target of two regions ensures the reliability of the senor in case one region undergoes no mutation. Furthermore, gold nanoparticles (AuNP) are coated with these single-stranded nucleic acids (ssDNA), which represent an ultra-sensitive sensor probe for the SARS-CoV-2 RNA.
The researchers previously showed the sensitivity of the developed sensor probes in their previous work published in ACS Nano. The hybridization of the viral RNA with these probes causes a change in the electrical response of the sensor. The AuNP shells accelerate the electron transfer, and when transmitted over the observation platform, it leads to an increase in the output signal and indicates the presence of the virus.
The team tested the performance of this sensor using COVID-19 positive and negative samples. The sensor showed a significant increase in the voltage of positive samples compared to the negative ones, and confirmed the presence of viral genetic material in less than five minutes. Furthermore, the sensor was able to distinguish viral RNA loads in these samples. Viral load is an important quantitative indication of the progression of infection and a challenge to measure using existing diagnostic methods.
This platform has far-reaching applications due to its portability and low cost. If the sensor is integrated with microcontrollers and LED screens or with a smartphone via Bluetooth or wifi, it can be used in the doctor’s office or even at home. In addition to COVID-19, the research team also predicts that the system will be adaptable for the detection of many different diseases.
“The unlimited potential of bio-engineering has always piqued my greatest interest with its innovative translation applications,” Alafeef said. ‘I’m glad to see that my research project has an impact on solving a real problem. Finally, I want my Ph.D. thank. Adviser Professor Dipanjan Pan for his endless support, research scientist Dr Parikshit Moitra, and research assistant Ketan Dighe for their help and contribution to the success of this study. ”
Reference: “Rapid, ultra-sensitive and quantitative detection of SARS-CoV-2 using antisense oligonucleotide-directed electrochemical biosensor chip” by Maha Alafeef, Ketan Dighe, Parikshit Moitra and Dipanjan Pan, 20 October 2020, ACS Nano.
DOI: 10.1021 / acsnano.0c06392