Abstract
We aim to design a point-of-care nitrite sensor coated with E. coli that has enhanced nitrite-digesting ability. This device can detect electric signals released from the reduction of nitrite and allows the public to monitor nitrite concentration in daily life.
There are only two simple steps to use our device. Firstly, the user adds the desired test sample on top of the sensor (pre-processed if the sample is solid). Secondly, the signal readout of the sensor will be sent to the mini-app on smartphone through Bluetooth with detailed analysis the and wait for the result.
Sensor Design
Our sensor could be generally divided into Arduino-based electrochemical detection system and screen-printed electrode sensing system. The Arduino-based detection system is based on the Gabriel N. et al.’ s tutorial in 2016 (Meloni, 2016). The nitrite sensor system is partially designed after Monteiro et al.’s research in 2015 (Monteiro, 2015), especially for the oxygen scavenger system.
Yet different from previous design, we choose to directly immobilize E. coli on the SPE surface to cut the cost and simplify the procedure to extract the nitrite reduction enzyme. Also, constructing engineered E. coli strains with high nitrite enzyme expression could improve detection efficiency and allows for continuous enzyme expression during detection. What’s more, immobilized cells help to maintain enzyme activity, therefore prolong the product lifetime and facilitate the storage.
Basically, the detection principle is to immobilize the enzyme (nirB & nrfA) containing E. coli on the SPE surface. Gelatin Methyacryloyl (GelMA), a commonly used hydrogel material, has been applied to coating for its high compatibility and low-cost (Heo, 2003). After using RIPA (Radioimmunoprecipitation Assay) lysis solution and GelMA lysis solution respectively for cell lysis and GelMA lysis, the nitrite reduction enzyme containing in the E. coli cells could be released form cell and absorbed to the porous carbon electrode surface. During the nitrite reduction enzymatic reaction, the nitrite will be reduced to the ammonium ion (NH4+) and have direct electron (e-) transfer with the electrode surface. The oxygen scavenger system is composed of gloucose, glucose oxidase (GOx) and Catalase (Cat) and is used to reduce the interference of oxygen in the detection reaction. Molecular oxygen (O2) is a main interferent in this analytical process because its reduction to hydrogen peroxide (H2O2) generates an intense cathodic current that can mask important redox processes that occur at very low potentials (Monteiro, 2019) . When applying cyclic voltammetry (CV) method for detection, there will be a linear relationship between the different nitrite concentrations in the detection sample and the current response in the detection circuit.
Figure 1: Scheme of electrode surface enzymatic reaction
Safety
At this stage, all nitrite reductases regulated by our transformed transcription factors used in our experiment need to be expressed in an anaerobic environment. On the one hand, it does increase difficulties in experiment implementation; on the other hand, it makes our bacteria safer to be storage and applied. Entrapping engineered bacteria in hydrogels further cut out the nutrient resources of the bacteria, preventing it from unexpected spread.
Application scenarios
We aim to design a point-of-care nitrite sensor coated with E. coli that has enhanced nitrite-digesting ability. This device can detect electric signals released from the reduction of nitrite and allows the public to monitor nitrite concentration in daily life.
There are only two simple steps to use our device. Firstly, the user adds the desired test sample on top of the sensor (pre-processed if the sample is solid). Secondly, the signal readout of the sensor will be sent to the mini-app on smartphone through Bluetooth with detailed analysis the and wait for the result.
Apart from the food, our product can also make a quick test of the water. It can be helpful for water inspectors to quickly get the nitrite level on-site before laboratory detection. Another typical example is the application in aquaculture where fish farmers will closely monitor the water nitrite level to avoid nitrite accumulation.
What is also worth mentioning here is that our devices allow the users to monitor the nitrite level of the plasma sample as well. As far as we concerned, the nitrite level in plasma could be related to many health issues such as endotoxemia, severe leptospirosis, and inflammation. Providing a point-of-care testing method could be essential in the early diagnosis of certain chronic diseases, which is especially important for special personnel such as the elderly and pregnant women. To draw a conclusion, as a new rapid test method, our product can not only promote safety awareness among the public but also help the aquaculture industries to minimize the harm of nitrite.
User manual
The following picture present how to use the “nitrisensor”
Figure 2-5
Software
In the end, the detected electric signal readout of our sensor could be read on the smartphone through Bluetooth. To visualize the result as specific concentration of nitrite, we developed an App associated with our nitrisensor. Thus, everyone could easily take control of their food security with some simple steps.
The App is also a platform for us to promote education with communities by publishing related popular science articles. Local news about recent nitrite-related pollution incidents is also available on the App.
Figure 6