The term "biosensor" is widely used for sensors that are based on biological sensing elements. With these, the targeted detection of specific substances (referred to as analytes) is possible. Biosensors have the potential to be used for a wide range of analytical purposes (Turner 2013). Furthermore, they can be cheaper, more sensitive and more specific than conventional detection methods like chemical sensors (Kaur et al. 2015). Consequently, biosensors are a field of active research and especially popular in the iGEM competition (Choffnes et al. 2011). Teams tried to use various different approach for implementing biosensors against toxic agents, hazardous chemicals, drugs, diseases and environmentally dangerous chemicals.

However, some issues remain: As great as the potentials of biosensors are, the hurdle of field applicability has in most cases not been overcome. This is because most biosensors make use of living, genetically modified microorganisms (GMOs) (Daunert et al. 2000, Lei et al. 2006). The application of these whole-cell biosensors is often not possible due to legal issues and safety concerns, as it must be prevented that genetically modified organisms are released into the environment. Moreover, these sensors are often not very user-friendly, as they have a limited shelf life and their application outside the laboratory is complicated and requires special equipment. Another disadvantage of using living cells as a biosensor is that the diffusion of the analyte across the cell membrane is a limiting step, which results in a slower response time of the biosensor (Strosnider 2003, Yagi 2007).

Nevertheless, there is a great need for handy and cheap detection tools for toxic substances, which became clear to us when we talked to experts and did a survey in Bielefeld. In our eyes, biosensors are very promising candidates for solving severe problems all over the world. Therefore, we decided to develop biosensors that overcome the aforementioned shortcomings and are easily applicable in everyday life.

After intensive research, we decided that such a biosensor needs to be cell-free. This has a number of advantages, especially in terms of biosafety and the legal situation. To make the biosensor easily usable for everyone, we aimed at developing a simple test strip. Paper has emerged as a cheap and reliable material for biosensors. By freeze-drying biological materials on paper, their shelf life can be extended significantly (Jokerst et al. 2012, Pardee et al. 2014). For this reason, it is the ideal material for our biosensor.

Another challenge in the field of biosensors is multiplexing (Chen and Rosen 2014). Often, it is not sufficient to test for one toxic substance to decide whether drinking water is safe or not, because most of the time several pollutants are present. Consequently, we wanted to build a modular biosensor that is easily extensible.

The basis of our biosensors is the interaction between repressor or activator protein and DNA. The principles are described in the following sections.

The repressor protein is a transcription factor which regulates the gene expression. It inhibits transcription upon binding the operator site of DNA, a DNA sequence for the transcription regulation. The complex formation can just be broken if the target substances (referred as “analyte” later) is available. They can bind the repressor which changes its conformation. The repressor protein is released and the transcription can start as the RNA polymerase is not hindered anymore.

The activator protein works on the opposite principle. It is also a transcription regulator and does not inhibit the transcription, but enables it. So by binding the analytes, the activator binds the operator sites. This time, the RNA polymerase can start to transcribe the genes as the affinity of the RNA polymerase to the promoter is enhanced.

In short: We decided to design a cell-free, paper-based test strip in order to make biosensors applicable in everyday life.

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