Overview
Here, we developed a G-quadruplex-based, highly specific, expedite and economic RNA virus detection system aiming to detecting the SARS-CoV-2 to halt the pandemic. On this page, we will demonstrate how we examined and subsequently implemented different measures to ensure the safety and security of all involved parties, from the development to utilization.
Considering Biosafety in Our Project
①Product safety - the saliva sampling: Currently, pharyngeal sampling and nasopharyngeal sampling are generally used in collecting specimens for COVID-19 tests. The two ways of sampling needs to put a swab close to the throat or reach the back of the naris, which can cause irritations leading to vomiting, coughing or sneezing. Thus, the saliva sampling, as a non-invasive collection method, is relatively safe as it protects the healthcare workers from being inadvertently exposed to the potentially infectious droplets. In addition to, sampling by pharyngeal or nasopharyngeal swabs has been associated with variable, inconsistent and false negative test results due to the technical difficulties of collecting proper specimens. Currently, the bottlenecks to improve accessibility and scalability of SARS-CoV-2 tests include diagnosis costs, complexity and supply chain shortages. To resolve these issues, we developed the SalivaDirect method. The critical component of our approach is to use saliva instead of respiratory swabs, which enables frequent non-invasive sampling and reduces the need of trained healthcare professionals during sample collection.
②Some nucleic acid detection methods typically require extraction of nucleic acid prior to amplification and multiple steps of manual operations, which undoubtedly complicate the testing procedure and potentially increase the risk of carry-over contaminations. However, in our approach, all components for nucleic acid amplification and CRISPR-based detection are thoroughly mixed in a single, one-pot reaction system, and incubated at a constant temperature, eliminating the need for pre-extraction and transfer of amplified products.
The Hardware
We used polypropylene to make the hardware, which is a colorless, odorless, non-toxic and translucent solid substance. Polypropylene is also a kind of excellent lightweight and universal thermoplastic synthetic resin. It also has great chemical resistance, heat resistance, electrical insulation, high strength mechanical properties and outstanding wear-resisting processing properties. For our overall reaction and subsequent digestion treatment, it provides the basis and assurance to guarantee a safe detection process and a stable detection system. We designed a "self-heating package" behind the color development step, so that the reaction mix in the hardware, including any remaining viruses and enzymes, is totally deactivated, to prevent any potential biohazard contamination leakage to the environment. Our system is dedicated to the safety of the environment and human body with the greatest extent to avoid any potential contamination to nature and human beings.
Experimental design: based on M13
The M13 phage has been considered as an attractive candidate in mimicking natural structures and developing revolutionary piezoelectric energy harvesting device. It only consumes bacteria, but is harmless to humans and environmentally friendly. It is also cheap and readily available, with trillions of the organisms harvested from infected bacteria in a middle-size flask. Importantly, due to the potential biohazard risk of SARS-CoV-2, it is unpractical to directly include the virus or any of its sequence in our study. Therefore, we used M13 phage to replace the COVID-19 specimen to verify the concept of our project.
