Team:SDSZ China/Demonstrate
Team:SDSZ China
To certify that our pollutant detection system is available in the real-world setting, we designed a kit for observation.
When infected by bacteriophages, the luciferase is released to the medium and would react with luciferin to produce fluorescent light. However, the light is not obvious when exposed to indoor lighting. Therefore, we made a sealable, light-proof box. The reaction tubes can be placed in the box, and as the box is sealed, the inside of the box is completely dark, and would be recorded by a camera placed in the kit. Therefore, the subtle fluorescence becomes highly palpable.
Besides the tube that contains our detection mix and water sample to be analyzed, we employed three control methods:
The first one involves two control tubes. Same amount of detection mix is added to each tube. In the first control tube, we add deionized water, the volume of which is the same as the water sample, to the detection mix; and in the second control tube, we add bacteriophage solution to the detection mix. With the positive and negative controls, we can exclude the false positive response and false negative responses caused by the detection mix itself.
The second one utilizes the Bl21 strain containing empty pET30a vector, cultured and induced under the same condition with the pET30a-luciferase strain. This provides another negative control that could exclude the influence of the strain or plasmid.
Moreover, as the cells would lyse within seven hours, we would control the reaction time within seven hours so as to prevent false positive response caused by cell lysis. With the control methods mentioned above, our detection results are more reliable.
Following the “Do Not Release” policy, we brought water samples from a domestic wastewater treatment plant, a sock factory sewage treatment plant and the fishbowl in one of our teammate’s home to the lab for testing. In the lab, we are able to put the reactants into our kit, and conduct our experiments without polluting the environment.
From the strength of the fluorescence, we can tell that the bacteriophage concentration is highest in the polluted water collected from the domestic wastewater treatment plant and lowest in the treated water collected from the sock factory sewage treatment plant.
After our detection, we put all the reaction mix under ultra violet light for 30 minutes, and then boiled all of them at 100℃ for 20 minutes so as to make sure that all the bacteria and phages are completely killed.
In the future, we would also integrate all these functions into our detection kit. The kit would contain two parts: the inner part is for reaction, containing our reaction tubes; the outer part is for safety, containing UV light and a heat source. In this way, our kit would be safe enough even for field tests. Concerning more safety issues, please turn to our safety page.