eLAMP: On-line viral diagnostic

We are committed to solve the problem of obsolescence/inadequacy of the current methods to detect infectious diseases. COVID-19 has crudely exhibited that the current standards of diagnostic, which depend on centralized labs, complex sampling, extraction and purification protocols, and highly trained personnel are not efficient to contain epidemic episodes.

We will complete the development and then pilot a portable device for the on-line, accurate, and robust diagnostic of infectious diseases from saliva in reduced time (i.e., three minutes). We will take COVID-19 as our first model.

This solution is designed to be portable, scalable, and amenable of self-use at home: an Arduino-based device that monitors, in real-time, an isothermal amplification reaction using a microelectrode. The massive scaling up of this solution will imply a change in paradigm, where on-line, quantitative, reliable self-diagnostic at home, and accurate location of positive cases and real-time monitoring will prepare us for the next pandemic. 

At present, more than 2.3 million people have been diagnosed with COVID-19 in México; SARS-CoV-2 has killed more than 240,000 citizens, making this the most lethal infectious disease in a century. COVID-19 has exhibit the severe limitations in our capacities to respond to epidemic emergencies and has made clear that we must expand and strengthen the portfolio of available tools for effective diagnostics of infectious diseases both now and in the future.

The retro-transcriptase quantitative polymerase chain reaction (RT-qPCR) is currently the gold standard methodology for COVID-19 detection. Despite its unquestionable accuracy and robustness, qPCR-based methods lack of portability, depend on centralized facilities, need for technical expertise to conduct the testing, and imply high infrastructural and operative costs. 

Due to all these limitations, RT-qPCR cannot be used massively in a cost-effective way to determine who is infected in a work place, at the entrance of an event, in a University classroom. We need robust, portable, cost-effective solutions that enable a safer comeback to our activities by quickly and reliably detecting even asymptomatic subjects. These solutions should allow fast diagnostic and localization of infected subjects (ideally in real time) to enable rapid decision making based on this information.

Our solution enables the portable, accurate, on-line, and cost-effective diagnostic of SARS-CoV-2 in any place (even self-testing at home).To do so, we integrate the use of two devices. Together, they render a completely electronic and quantitative device that can test and report results on-line for real-time diagnostic and precise location of positive cases. 

First, we use a paper-based microfluidic system (made from ultrafiltration membranes and lamination film) for the isolation and concentration of SARS-CoV-2 nucleic acids from saliva. This paper-based device absorbs the liquid in the saliva and retains the solids in a small area (a circle of 5mm in diameter). This circle is cut and transferred to the second device that consists in a compact Arduino-based incubator that can be hold in the palm of your hand. If present, the SARS-CoV-2 genetic material is isothermally amplified in this incubator using a small volume of reaction mix. The electronic system enables the on-line monitoring of the amplification by using a pH microelectrode, and displays the change of electric potential in the reaction mix associated to the amplification. A communication system will  enable the real-time transmission of the amplification curve (the result) and the precise location where the test was conducted. 

Our solution serves individuals (i.e., by providing means of reliable, portable, and cost-effective diagnostics of COVID-19; extensible to other infectious disease). Also, our solutions serves the well-being of the community by (a) enabling a more frequent and simpler testing of individuals and the detection of cases (inclusively asymptomatic) and (b) by providing a strategy for frequent and fast testing at any office, any workplace, any school, and even every home. 

The design of an interface capable of collecting in real-time all the data from all the curves of amplification of all users of this testing device will allow us to elevate the impact of eLAMP. We believe that our solution will be greatly helpful to monitor, in real-time, the number of cases and their regional distribution within a community. The data collected using this new technology will enable the easier identification of zones of high incidence or high risk. That cannot be easily done (at least in Mexico) using data derived from RT-PCR. 

The data collected in practically real time from all tests conducted using eLAMP will also serve the needs of decision makers. Much faster response will be possible to epidemic onsets in particular locations (any school or working place) by monitoring the number and trend of positive cases over time. 

During pandemic COVID-19 we have seen that the implementation of widespread diagnostics and selective quarantine can effectively buffer epidemic curves. Korea, Germany and other European countries have set the example in this regard. Numerous research papers have described how opportune widespread diagnose is key to contain pandemic events. We recently published a paper that quantifies the effect of different diagnostic intensities in containing local onsets of COVID-19 (Alvarez, M. M., González-González, E. (2021). Modeling COVID-19 epidemics in an Excel spreadsheet to enable first-hand accurate predictions of the pandemic evolution in urban areas. Scientific reports, 11(1), 1-12.). Our simulations clearly show that opportune and widespread diagnostic is highly effective. The effectivity of the ongoing worldwide vaccination programs will be highly enhanced if they are combined with aggressive diagnostic programs. (https://www.medrxiv.org/conten...).

Our solution will be of great value to our society because it enable both faster and accurate diagnostic of every individual. In addition, our technology is portable and easily deployable. Also, we use saliva to detect SARS-CoV-2 which is much less intrusive than using nasal or pharyngeal samples (the current standard). Currently we do not have a technology that combines this attributes to respond to this and future pandemics. 

COVID-19 showed to us all the limitations of RT-PCR from nasal/pharyngeal samples, the current gold standard for the diagnostic of infectious diseases: complex (and uncomfortable sampling), complicated extraction and purification protocols, dependence of costly and non-portable lab equipment, and throughput limitations. 

Our solution will enable portable, accurate, on-line, and cost-effective diagnostic of infective diseases in any place (even self-testing at home).

Then, our solution fully aligns to one of the axis of this challenge: preparedness for a quick response in pandemic episodes. Precisely, it is an early detection system that detects future disease outbreaks in real time strengthening disease surveillance. 

 We have developed a very straightforward strategy to concentrate nucleic acids from saliva samples using a laminated segment of PES-membrane. In addition, we have a (proof-of-principle) working Arduino-based prototype that is capable of discriminating between saliva samples containing or not containing synthetic genetic material from SARS-CoV-2. The next stage for our solution is to miniaturize, to optimize, and to present these two components in a single, coherent, and streamed system. Then, we aim to develop the hardware and software required for our platform to transmit the amplification curves to the cloud as measured (in-real time). 

Integrating and packaging all these features will yield a prototype ready for validation and eventual approval from regulatory and public health authorities at Mexico.

Most platforms for viral diagnostic depend on the use of nasopharyngeal samples and depend on centralized labs. Point of care diagnostic systems have been implemented experimentally, but most of them are qualitative. Our solution is compact, portable and reliable. This makes it amenable for massive implementation and suitable for self-diagnostic, even at home. 

This is disruptive, and may change the standard of testing. Currently, the patients send samples to a centralized facility for testing. In the future, systems such as ours may enable distributing kits to the location where the patient is (i.e., using a service such as Amazon or Uber). Patients will be able to do self-diagnostic. Moreover, our system is fast and completely electronic, which enables quantitative on-line diagnostic. The result and the location where testing was done can be sent to the cloud in real time, which is both highly useful and revolutionary.   

This will enable a truly real-time monitoring of the pandemic, which can not be done today.

We are currently developing the first functional prototype of our system. As part of the initial characterization of the performance of our first prototypes, we have (presumptively) diagnosed approximately 30 individuals. We envision that our solution will serve between 10,000 and 100,000 individuals in the next year. 

Our solution has the potential of serving tens of millions of individuals within the following five years. We address cost-effective and portable diagnostic of infectious diseases taking SARS-CoV-2 as a model; the number of tests that were administered worldwide during this last years must be in the order of billions. Our solution can be easily adapted, by changing the sequence of the primer sets, to identify in saliva any other infectious disease that could be transmitted by aerosols. 

We have mainly evaluated our progress in terms of the performance of the prototypes that we have produced. Sensibility (ability to correctly identify positives) and Specificity (ability to correctly identify negatives) are the two main indicators used to evaluate the accuracy of a diagnostic tool. The comparison is made against a reference method (i.e., for real samples the current accepted gold standard is RT-PCR). The third indicator that has been followed during development is time to get a result.

Our first prototype exhibited good sensibility (95%) but not equally good selectivity (aprox. 85%). A second version of LAMP that derived from our research is being piloted by Aim Brands in USA. Using this pre-commercial method we have observed very good sensibilities (95%) and improved selectivity (aprox. 92 %). Time to get results with our current system is 40 minutes. Our preliminary data based on synthetic saliva samples suggests that eLAMP exhibits sensibilities and specificities near 100% in less than 5 minutes. We expect to see a similar scenario in real saliva samples. 

As implementation in the community begins, the number of tests conducted (people diagnosed) and the number of organizations adopting this solution will become equally important indicators of success. 

We are the Alvarez-Trujillo Lab, a research group within Tecnológico de Monterrey (a non-profit private University in México). Our lab currently has more than 40 active members. Two faculty members, one posdoctoral researcher, two pH.D. and one M.Sc. student, and two undergraduate students will be directly involved in the project.

Our group has been working in diagnostic systems since the times of pandemic Influenza A/H1N1/2009, which started in Mexico. We have been working in the components of this solution since March 2021 (15 months).

One of our main strengths is our multi-disciplinary nature. Our laboratory integrates professionals from many different disciplines, majors, and/or profesional experiences from molecular biology to hardcore engineering (chemical, electrical, mechanical engineering, mechatronics, nanotechnology).

In addition, we have people trained in México, Spain, England, India, Italy, Switzerland, Iran, and USA. Some of our students and postdocs have family and friends in all of these countries and relate well with the way people in different regions have lived throughout pandemic COVID-19. The leaders of the team, Grissel Trujillo Mario Alvarez have posdoctoral experience at MIT and Harvard. 

Lastly , we have been very active as a team during these months. A list of our contributions in the front of COVID-research follows:

• Alvarez, et al. 2021. Portable and label-free quantitative Loop-mediated Isothermal Amplification (qLAMP) for reliable COVID-19 diagnostics in 3 minutes: An Arduino-based detection system assisted by a pH microelectrode. medRxiv.doi: https://doi-org.ezproxyberklee.flo.org/10.1101/2021.0...
• González-González, et al. 2021. Colorimetric loop-mediated isothermal amplification (LAMP) for cost-effective and quantitative detection of SARS-CoV-2: the change in color in LAMP-based assays quantitatively correlates with viral copy number. Analytical Methods. 13(2):169-178.
• González-González et al. 2020. PLoS ONE 15(8): e0237418.
• Marquez-Ipiña AR, et al. 2020. Diagnostics. 11(2):217
• Alvarez, et al. 2020. Scientific Reports. 11 (1): 1-12.
• Mahlknecht, 2021. Science of The Total Environment, 147183.

As a research group, we are committed with diversity and equity. The current top leadership of our group is shared between Grissel Trujillo de Santiago and Mario Moisés Alvarez. Currently, both of our posdoctoral researchers are women (a Mexican trained in Spain and a German trained in Switzerland and England). We currently have students from México, Colombia, USA, and Iran. These information would probably not be surprising in an international environment of entrepreneurship or technology development. However, Mexico is still evolving to be an inclusive and diverse society, and for us is truly an honor and achievement to be particularly diverse in the Mexican context. Proudly, our research group is integrated by more than 60% of women. In México, that is high for a research group working in Engineering.  

The main value that we are aiming for is exposure and dissemination of our idea. We truly believe that our solution has the potential to dramatically change the current paradigm of diagnostic based on centralized labs. 

In Mexico, where we are currently located, pandemic COVID-19 exhibited all the limitations of the current state of the art in diagnostics. We strongly envision the contribution of our solution in preventing that this happens again in Mexico or in any other place, in the context of COVID-19 or  any other emergent infectious disease. 

Of course, we are also in need of economic backup. We have been able to obtained a reduced amount of funding (nearly 10,000 USD) to produce a functional prototype. Validating and piloting this solution In Mexico has been difficult due to the position adopted by the Mexican government in the sense of only adopting and implementing "validated technologies" based in the analysis of nasal or pharyngeal swabs for molecular testing. We strongly believe that our straightforward technology should be tested/piloted in other countries to accelerate massive adoption. In that sense, Solve is an ideal platform were our idea can reach the correct audience. This call is particularly suited for the solution that we are proposing. 

We believe that our solution can dramatically change the way we conduct diagnostic rights now (mainly in centralized labs). We have very clear that we can do it, but we need more people to see this feasible. In other words, we need to showcase our idea and our prototypes. Our solution is so simple, so obvious, that can be easily obscured by more sophisticated and complex ideas. 

We believe that SOLVE opens many extraordinary possibilities of presenting our ideas and our solution to experts, investors, scientists, decision makers around the globe. Then, we are mainly interested in the network that SOLVE can give as access to, more than the funding.

At this point, the money is a valuable resource However, what really will take our solution to the next level is really the networking and the visibility that SOLVE can give to our project. Of course, we are in great need of that the partner that believes in the idea and decides to pilot it somewhere.

We envision partnerships around the globe with organizations, governments, universities, that could pilot the use of our diagnostic system in open population. We have run hundreds of validation assays with synthetic samples of SARS-CoV-2 genetic material. However, we need to give the following step and validate our solution with real samples in controlled experiments, and then in the field. 

We are also in need of partners that could help us to package this technology in the form of a final and robust product. Evidently, we are open to partner with investors, start ups that could see the potential that we see in this solution to really change the way we conduct the diagnostic of infectious diseases.

Our solution aligns perfectly well to the aims of the Robert Wood Johnson Foundation: elevating the health and well-being of everybody in the USA. We believe that COVID-19 pandemics clearly demonstrated that the unprivileged is the one that suffers the most during an epidemic emergency. Different studies showed that in the USA and around the world. 

We believe that our solution can reach any corner of USA in a cost-efficient manner. We envision that the commercialization of our diagnostic device (a controlled temperature incubator capable of discriminate between positive and negative subjects in less than 3 minutes through on-line readings of electrical potential) is feasible at $200-300 USD per device. The cost of the reagents may be as low as $3 dlls per sample. Tese numbers enable the many possibilities to reach unprivileged sectors of the population. For example, we envision mobile units capable of do at-home diagnostic in unprivileged areas. Self-diagnostic at home is a valid possibility to reach any zone in a urban area using the United States Postal Service, or private services such as Uber or Amazon. 

We anticipate that our solution can be of great use for Refugee populations. In a moment of an epidemic crisis, refugees are one of the most vulnerable populations, with limited access to diagnostic and health services, in general. In parallel to pandemic COVID-19, we observed a refugee/migrant crisis in the Guatemala/México border and the Mexican/USA border. The characteristics of our diagnostic solution (portability, accuracy, use of saliva samples, quick response) perfectly fit the basic requirements for implementation in refugee shelters and immigrant communities. 

Our solution may be of great value to continuously diagnose health workers and volunteers while they support communities. Three characteristics of our diagnostic solution allow frequent use everywhere. Testing is done from saliva, which greatly facilitates frequent use. Portability enables reaching remote communities. Diagnostic devices can become part of the equipment that volunteers and health workers could carry in their backpacks. The possibility of self-diagnostic reduces risks of infection among health workers and volunteers. Conceivably, each person can self-diagnose minimizing the risk of handling infective samples from one place to another.