Overview
Our Project
Our project aims to provide a fast-reaction home test kit for liver diseases by qualitative analysis of urine bile acids levels. Considering the inconvenience and high cost of hospital testing, we use synthetic biology to design gene circuits and construct the biosensor with three devices: substrate processing device, detection device and report device. Meanwhile, we try to expand our project into a test kit with corresponding hardware.
Our Final Product
The final product of our team is a home test kit which uses bile acids levels as the indicator to diagnose liver diseases. The lyophilized content of the kit is separated into two tubes A and B. The tube A includes the substrate processing device and the tube B is for detecting and reporting. Users only need to inject water into the tubes at night before using, so that the cell-free system can be incubated to a sufficient concentration of proteins to hydrolyze the 3-sulfate esters of bile acids. On the next day, users can collect urina sanguinis, and dilute it 5 times in tube A and wait for 2 hours. Then they just need to pour the contents of A into B and wait for 8 hours. The color of tube B activated by the light-emitting device will tell users if they are at risk of liver diseases. Our kit has been proven to achieve our expectations at room temperature.
Kit Instructions
Device
The complete test kit includes a measuring cup, a dilution device and a detection device.
Measuring Cup: A container with a fixed scale for urine.
Dilution device: The device includes a diluting vessel and diluting solution (ddH2O).
Detection device: The device consists of a hand-held fluorescent illuminator as mentioned in the literature, a cell-free sensor (here we use lyophilized powder) designed and further developed in this project, and a container that resembles the one provided in the literature, surrounded by an outer casing (as shown below in Figure.1), and above it lying a channel for the urine sample to be tested.
Figure 1. The hardware
Steps
Since qualitative diagnosis requires a fixed dilution ratio, users need to dilute their urine sample initially. Firstly, users are supposed to take a certain amount of urine in the measuring cup. Then, they need to dilute the urine step by step by pouring it into the diluter. Finally, after the diluted urine sample being poured into the detection device, users can turn on the light-emitting device and observe the color change of the receptor.
After a certain period of time (the set incubation time), if the sensor shows an obvious red color, the user may need to seek immediate medical attention for liver diseases. Whereas the case that the sensor doesn’t show any evident color means medical attention is not necessarily required.
Storage
Storage at 4°C in Refrigerator (for home use).
The Further Development of Our Project
Apart from the development of test kit, we also envisioned a Liver Nursing WeChat applet as its subsidiary service. We listed several principal needs of chronic hepatitis users as below:
1. Nursing needs: Generally speaking, chronic liver diseases require long-term treatment, reasonable diet and lifestyle management. Unfortunately, most patients lack recordings and management of their life.
2. Psychological needs: Some patients may suffer from greater psychological pressure after being diagnosed with diseases. Studies have shown that reasonable psychological nursing intervention for chronic hepatitis patients is of great value to their psychological state and disease recovery.
3. Educational needs: Most families have little knowledge about chronic hepatitis. If their awareness could be raised, it would be of great help to improve their situation from their living habits and play a preventive role.
Eventually, based on extensive investigations and interviews, we implemented the idea of establishing a WeChat applet of liver care for our potential users. We offered the following functions for the platform:
1. Life Diary
To help clients better manage their lifestyle, this section is used to record their diet, sleep, mood, and exercise. There are 4 corresponding recording areas. After the selection of this module, one can write down his/her life in these 4 dimensions and the corresponding data will be compared with our preset healthy lifestyles, then it will be evaluated and corresponding personalized health advice would be formulated and provided to clients automatically. Actually, there are already numerous mature programs for this design on the market. We learned about outsourcing services in the previous communication with the programmer Mr. Qi, so we hope to seek help from some experienced IT companies in the future. We are also planning to add recommended recipes and timetables for hepatitis patients in this module currently.
2. Hepatitis Mini Class
To improve people's awareness of hepatitis, this section provides short video lessons and related articles on hepatitis. During the early-stage investigation, it was found that the public generally lacked the understanding of the occurrence and development of hepatitis, and the current level of knowledge and education about hepatitis in China was not enough either. Thus, we intend to help people make the most of their fragmented time to learn about liver health through lightweight knowledge education short videos. We will keep on uploading high-quality videos and articles so that people can gain more information about liver health in this module in the future.
3. One-to-one oriented psychological service community
To meet people's communication needs regarding liver disease and relieve the psychological pressure of chronic hepatitis patients, this module aimed to provide special communication services to our clients. Studies have shown that reasonable nursing intervention for chronic hepatitis patients is of great value to their mental state and recovery. So we decided to provide psychological counseling services. We realized that although the resources of professional doctors are in short supply, the huge network of customer service resources in hospitals is currently in a state of oversupply. Most of those staffs in medical network services are well-trained and have sufficient knowledge of many diseases. They mainly provide disease consultation and online communication services. Actually, they can fully meet the needs of patients instead of turning to doctors in business. Plus, the hospital network customer service is not equal to the hospital hotline customer service, and our services will not occupy the hospital's hotline at all. Based on this discovery, we integrated the hospital network customer service into our applet. Patients could initiate an online discussion in this section, and they could be automatically connected to the hospital network customer service for communication.
Concerning Safety
For Products
Our final product kit contains lyophilized powder, which may cause related biosafety issues due to the leakage of the lyophilized powder into the environment. Therefore, in possible future applications, we strongly recommend that our users avoid damaging the test kit when using it, and hand over the used kit to professionals for recycling.
For Experiment
Throughout the whole experiments, we have fully complied with iGEM's security policy. All our validations were done in the laboratory without releasing any engineered bacteria into the environment.
In the implementation stage, the cell-free sensor used in the pilot scale test was consistent with the prototype in our laboratory. When producing cell-free sensors in the form of lyophilized powder, we handled them in strict accordance with regulations to avoid possible biosafety issues.
Challenge to Conquer
Application of kit
1. The composition of urine is comparatively complex, generally including urea, creatinine, a small amount of protein, etc. Besides, the pH value also varies greatly. As a result, related tests are needed to take the effects of urine composition and properties into account.
2. Due to safety concerns, we could not obtain urine samples. So we are hoping that relevant qualified and licensed institutions can help us with more real-world investigations.
3. It is reported that there are differences in the content of urine CDCA of people in different regions. Therefore, when applying the kit in different regions, it is necessary to take the actual situation into consideration and conduct extensive investigations by relevant personnel before further application.
4. Further exploration of the necessary application details remains being discovered, such as the storage time of the kit, the maximum endurance of the kit, the amount of dry powder contained in each detection device, etc.
Reference
Jung, J.K., Alam, K.K., Verosloff, M.S. "Cell-free biosensors for rapid detection of water contaminants". Nature Biotechnology 38.12 (2020):1451-1459.
Tu, Hua, Arthur Y. Okamoto, and Bei Shan. "FXR, a bile acid receptor and biological sensor." Trends in cardiovascular medicine 10.1 (2000): 30-35.
Alford SC, Abdelfattah AS, Ding Y, Campbell RE." A fluorogenic red fluorescent protein heterodimer". Cell Chemical Biology 19.3 (2012):353-60.
Makishima, Makoto, et al. "Identification of a nuclear receptor for bile acids." Science 284.5418 (1999): 1362-1365
Foreman, Kyle J., et al. "Forecasting life expectancy, years of life lost, and all-cause and cause-specific mortality for 250 causes of death: reference and alternative scenarios for 2016-40 for 195 countries and territories." The Lancet 392.10159 (2018): 2052-2090.
Moon, Andrew M., Amit G. Singal, and Elliot B. Tapper. "Contemporary epidemiology of chronic liver disease and cirrhosis." Clinical Gastroenterology and Hepatology 18.12 (2020): 2650-2666.
