Aalto-Helsinki Team Wiki

Aalto-Helsinki Team Wiki



Zooming Into GutLux

To better understand GutLux, we can break it into two different levels:

  1. The Capsule
  2. The Compartments

To understand the evolution of our project and how we decided on building an ingestible biosensor, check out our Human Practices page.

The Capsule

GutLux is an encapsulated biosensor with internal detective components. The GutLux capsule allows gut material to enter the capsule for detection within the gut. It is pill-shaped so it can be easily ingested by the user -- like a vitamin pill. While building the capsule, several factors were taken into account such as gut conditions like the pH levels, the shape and structural composition of the intestines, and the physical forces found within a functioning digestive tract.

With these factors in mind, we designed our capsule with specific biomaterials compatible with the gut, and engineered it to be of a particular shape to not only make ingestion easier, but to enhance its safety and efficacy within the digestive tract. To learn more about the considered biomaterials, please refer to our Hardware page.

Given the shear forces found with the peristaltic movements in the gastrointestinal tract and the winding path our capsule would travel through before its exit, we anticipated some potential safety concerns. Namely, the potential for the capsule to break or rupture in vivo, or for its shape to cause harm to the soft tissues of the gut wall. We have designed the capsule with rounded corners so that there are no sharp edges that can scratch or puncture the gut epithelium. GutLux has also been shaped to be structurally resistant to damage, so that in its movement through the GI tract, forces that propagate the bolus forward will not easily break the capsule. For more information on GutLux shape and dimensions, click here.

To ensure the capsule’s efficacy within the gut, we understood that it would need to be engineered to withstand the harsh conditions, notably the varying pH levels. GutLux encounters areas of the GI system with a pH range of 1.0-7.5 (Evans et al., 1988) prior to reaching the desired detection area. Therefore, we are planning on including a pH sensitive covering that will protect it before it reaches its destination --the intestines.

To allow for metabolite detection, GutLux contains a semipermeable membrane that will allow for luminal contents including our metabolite of interest to enter the capsule.

The Compartments

GutLux can be broken down into two compartments to facilitate three different functions. The first compartment contains our biological system, which in the presence of a metabolite, facilitates the biologically-mediated production of fluorescent light in response to the metabolite’s concentrations. The second compartment contains the electrical components which are responsible for measuring the intensity of the fluorescent light signal produced, converting the measurement to digital data and then transmitting the resulting electrical signal to an external device.

The Biological Compartment: From Metabolite Concentration to Light Signal

GutLux’s detection mechanism occurs through the human aryl hydrocarbon receptor - otherwise termed as AhR. This receptor has been well characterized to bind to a variety of substrates, some of which are of interest to our project. The mechanism functions with a trio of proteins: the human Aryl Hydrocarbon Receptor (AhR), the Aryl Hydrocarbon Receptor Nuclear Translocator (ARNT) and the AhR Interacting Protein (AIP). Together, these proteins function to produce a downstream fluorescent signal once bound to our metabolite. Our protein triad is expressed in a single plasmid and upon ligand binding, interacts with a separate reporter plasmid that will produce the fluorescent protein within the whole cell. Our detection system is visualised in Video 1.

Video 1. Biological detection mechanism.

Choice of Metabolite

With our biosensor, we aim to better understand the connections between our gut and our mental health. In order to do so, we planned to measure the concentrations of metabolites that have been preliminary associated with mental health conditions. More specifically, we were interested in looking at metabolites related to tryptophan, as tryptophan and its derivatives have been linked to depression (Averina et al., 2020), anxiety (Funakoshi et al., 2011), and bipolar disorder (Sellgren et al., 2019). Given the research nature of GutLux, we hope to increase the understanding of our chosen metabolites and their general impact on gut and mental health overall.

Tryptophan is metabolized in one of three pathways: the serotonin, kynurenine and indole pathways. The products of these pathways are shown in Figure 1. Within the list of these derivatives, we decided to focus on kynurenic acid and tryptamine.

Figure 1. Tryptophan and its metabolism pathways (Roth et al., 2021).

Kynurenic Acid

Kynurenic acid is a tryptophan metabolite that has been reported to exhibit neuroactive activity when it is of neuronal origin (Reyes Ocampo et al., 2014). It is derived from kynurenine in the kynurenic pathway, and has previously been reported to be present additionally in the distal small intestine and the proximal colon (Turski et al., 2013). The origins of said kynurenic acid have been reported to come from a variety of sources. For example, it has been reported to be present in human gastric juice leading to the assumption of endemic kynurenic acid production (Paluszkiewicz et al., 2009), as well as the liberation of kynurenic acid through aspartate aminotransferase activity inhabited by gut microbiota Escherichia coli strains. Higher concentrations were also reported in rats when looking at potential anti-depressive properties of certain probiotics (Desbonnet et al., 2009).


Tryptamine is an indolamine tryptophan derivative that is created through tryptophan decarboxylase activity in the indole pathway branch. This decarboxylase activity in humans is only possible through two bacterial species, Ruminococcus gravus and Clostridium sporogenes, both of which inhabit the human gut microbiota (Williams et al., 2014). Tryptamine is normally found in plants, but these bacterial species responsible for the creation of tryptamine have been reported to be present in 10% of adults (Williams et al., 2014). If present, it is found exclusively in the gut lumen, as it is dependent on microbiota reactions for its synthesis. It has been reported to induce the release of serotonin from the enterochromaffin cells in the gut (Roager, Licht, 2018). Studying tryptamine concentrations will not only help researchers better understand any potential roles of tryptamine, but also its interaction with serotonin release and its downstream effects.

In addition, GutLux aims to fill a gap in diagnostic testing by observing a completely different sample type -- real time gut contents. Unlike traditional diagnostic samples such as blood, feces and urine, GutLux aims to observe what is happening in the gut in real-time, without interference and alterations associated with sample collection outside of the body. As such, our choice of metabolite had to be specific to the gut, and most importantly, must be present within the gut lumen. Since GutLux will only be present within the lumen, and will not enter the gut epithelia, some potential metabolites were excluded, for example, serotonin. Serotonin has direct reported links to a myriad of mental illnesses, but since gut serotonin is synthesized on the apical side of the gut, GutLux would not have access to it.

Type of Biosensor

To detect tryptamine and kynurenic acid, we had the option of choosing to create either a molecular biosensor, a cell-free biosensor or a whole cell biosensor.

When looking at potential molecular biosensors, we looked at mechanisms that would generate a readable signal from metabolite binding through a variety of enzymatic reactions. Albeit there being a few options for our interested metabolites, we found that they would severely complicate the detection mechanism and introduce a lot of room for error. For example, one promising mechanism used glucose as an important downstream factor after metabolite binding. We were almost certain that luminal glucose would enter the capsule, of which would create a false positive signal. We would no longer be able to equate the signal being produced to represent the amount of metabolite present. Other enzymatic reactions included multiple steps which we anticipated would result in a high probability of incomplete reactions.

After determining a protein cascade that would detect our metabolites, we looked at systems that supported this. The AhR system was particularly favourable because it is found naturally in humans and is capable of binding to a variety of tryptophan derivatives. When considering a cell-free system, albeit its benefits (i.e., reducing the ingestion of genetically modified organisms (GMOs)), we determined that it would not be feasible given the nature of our biosensor. Since our biosensor has somewhat of an open “window” that allows luminal contents to enter at any time, and the fact that molecule concentrations change throughout the digestive tract, we could not ensure that the necessary components of the cell-free system would not leak out. Current membrane technologies separate based on size or charge, lacking the specificity needed to prevent components from leaking out while allowing metabolites in. As such, we anticipated small, necessary components such as adenosine triphosphate (ATP) and redox factors to leak out of our capsule, and harm GutLux’s ability to detect metabolite concentrations accurately.

With these factors in mind, we decided on using a whole cell biosensor to facilitate our detection mechanism. We would be able to produce all the necessary proteins for detection within a model organism and couple it with a fluorescence production system described below. Some additional benefits included the cost effectiveness and the resourcefulness of a whole-cell biosensor as there is no need to purify proteins and our cells would self replicate.

Choice of Microorganism

Our detection mechanism has been adapted in both Escherichia coli and Saccharomyces cerevisiae. Since our detection protein is characterized in humans, we wanted to observe how the system would perform in both prokaryotic and eukaryotic cells.

Given our access to E. coli strains, its widespread uses in synthetic biology and its overall reputation as a safe, well-characterized model organism, we decided to use it as one of our microorganisms. More specifically, we decided to work with two different strains: TOP10 and C2566. TOP10 was used when performing transformations as it helps with increasing transformation yield. Alternatively, C2566 is a BL21-derived strain, and was used as the chassis in which our three proteins would be exposed to our green fluorescent protein-containing plasmid. When working with E. coli, we expressed truncated versions of the AhR and ARNT proteins. This was due to the fact that previous researchers have had concerns with expressing full-length versions of AhR and ARNT in E. coli, but truncated versions were expressed successfully in this article (Schulte et al., 2017).

When choosing our model eukaryotic organism, one of our main priorities was its life cycle duration, given the fact that eukaryotic organisms take longer to work with in comparison to prokaryotes and we had a limited time frame. We chose S. cerevisiae for its shorter turnaround time in comparison to human cells, its characterized safe uses within humans and most importantly, the reported successful in vitro expression of full length AhR (Miller, 1997). More specifically, we used the W303ɑ strain as it contains silenced copies of marker genes that would help facilitate the necessary integration of our gene fragment of interest into the S. cerevisiae genome and also allowed for the successful selection of recombinants.

For further information on how the planning of our experiments varied between the two organisms, click here and how our workflow differed between the two, click here.

The Electrical Compartment: From Light Signal to Electrical Signal

We opted to have detected metabolite concentrations represented by fluorescent light. When deciding what signal we wanted to measure, we looked at a few options. Fluorescence was always a strong contender, as fluorescent proteins within reporter systems were readily available and it has been used countlessly in research, even in biosensors similar to ours. We also considered luminescence via luciferase which would require the addition of an additional substrate. Given that the capsule is ingestible, we would not be able to supplement the capsule with additional substrate required for the luminescent reaction should it run out. As such, we discarded this option. One completely different method of quantifying compound concentrations measured the electron transfer through an electrode in an enzymatic reaction that would occur after the detection of tryptamine. Albeit an interesting method, we excluded this idea as we couldn’t resolve the safety concerns regarding the ingestion of graphite and gold nanoparticles and the production of toxic H2O2 that would be essential in this method.

After deciding on using a green fluorescent protein as our source of fluorescence, we worked to optimize the detection of this signal. To learn about how we did this, refer to the Sensing section on our Hardware page.

The Electrical Compartment: From Electrical Signal Transmittance to External Device

After our fluorescent signal has been measured, it is converted to an electrical signal and then transmitted to an external receiver that contains the data collected which can be later analyzed. GutLux’s electrical signal will be transmitted to the device via radio frequency. There were a myriad of potential options for transmittance such as Wi-Fi or Bluetooth, but given the ingestible nature, the ethics of data privacy, the distance between the capsule and the receiver and the proposed end-users, we decided on radio transmittance. To learn about specific frequencies we use and the transmitters and receivers we chose, please visit the Transmittance section on the Hardware page.

Furthermore, we looked at the safety and privacy concerns of transmittance within the gut. We hosted a Data Privacy and Security workshop alongside our partnering iGEM team, iGEM Team Thessaly 2021, where we discussed the reasoning for our different chosen methods for transmittance despite both teams building an ingestible biosensor. To read about this workshop, please refer to the Partnership page.


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