We designed the capsule with four circular main electronic printable circuit boards (PCBs) with light-emitting diodes (LEDs) and light-dependent resistor (LDR) board, processor board, radiometry board and power module. This design optimizes the used space within the capsule and enables the small round shape. The PCBs are stacked and connected with headers. Our four layer PCB design is illustrated in Figure 1. Similar designs, with stacked round PCBs, were used for an ingestible capsule that measures gas composition inside the gut (Steiger et al., 2019) and for an ingestible capsule that also utilizes LEDs and LDRs to measure fluorescence and detect colorectal cancer (Alam et al., 2020).
The capsule dimension and shape were chosen so that risk of retention is minimized. Capsules smaller than 11 mm in diameter and 28 mm in length are regarded as 100% safe (Kalantar-Zadeh et al., 2017). The pill-shaped form of the biosensor supports the ease of ingestion, and the capsule has been designed with rounded corners so that there are no sharp edges that can scratch or puncture the gut epithelium.
To measure the concentration of expressed green fluorescent protein (GFP), a sensing board for the 4 layer PCB was designed. The GFP needs to be excited for it to emit fluorescence, so we designed the sensing board similar to what Alam et al. (2020) created. The sensing board consists of LEDs, which excite the GFP, and LDR that can then measure the change in light intensity. Our biological detection mechanism utilizes the superfolder green fluorescent protein (sfGFP). From Figure 2, we can see that the emission peak of the sfGFP is 485 nm and the excitation peak is 510 nm. Since the excitation and emission overlap, we might need to use light filters in the light pathway to block specific wavelengths.
During our project development, we realized that it would be beneficial to have a product that could utilize multiple fluorescent proteins and could maybe even detect multiple different wavelengths if we wanted to detect many different metabolites simultaneously with one biosensor. Hence, we wanted to develop a system that can later be altered to detect other wavelengths. For LDR, we chose to use multichannel surface mount LDR. This would allow us to use multiple fluorescent proteins and optimize the detection wavelength. The surface mount component that we chose is AS7341, 11-Channel Spectral Color Sensor with dimensions of 3.1 mm x 2 mm x 1 mm and has a channel with center wavelength of 480 nm.
The LDR is situated in the middle of the PCB with four LEDs surrounding them. The chosen LEDs for this application should be small, so for our surface mount LEDs, we chose LXZ1-PE01 with 500 nm center wavelength and size of 1.7 mm x 1.3 mm x 0.66 mm.
To further communicate the measured light intensity from the capsule to the outside receiver, we need to transmit the data through wireless transmission technology. We decided to use radio frequency (RF) transmittance. This technology is already established for ingestible sensors and so, there is a considerable amount of research of RF transmittance for ingestible sensors (Nikolayev et al., 2017). But implementing this method needs a lot of consideration, so that the quality of the transmission meets the needs. After discussing with Kaisa Linderborg, associate professor in molecular food sciences with experience in using ingestible capsules for research, we learned that one of the main problems with current capsules is the connection breaks between the ingestible sensor and receiver. This leads to gapping in the data and data losses. Indeed, sending data from inside the body is challenging, since surface interference and signal deflection interrupt the signal (Vasisht et al., 2018).
To tackle this issue, we looked at a few different communication bands used for telecommunication for ingestible capsules. In Given Imaging capsules, the used communication band is 433 MHz (Kalantar-Zadeh et al., 2017). It is mentioned to be one of the most suitable for this purpose, since the signal passes through the body with low propagation loss. According to Nikolayev (2017), the band between 433 to 434.8 MHz is the most suitable, unlicensed and globally available range of frequency bands for in-body devices. There are also studies that suggest that transmission frequency of 400-900 MHz would be optimal. Since the RF signal of longer wavelengths propagates increasingly in body tissues, we chose to focus RF transmitters and receivers with a communication band of approximately 433 to 434.8 MHz (Steiger et al., 2019).
For in-body communication, far-field RF signals are the most suitable, since they radiate omnidirectionally from the device and thus allow multiple orientations of the capsule in the gut (Steiger et al., 2019). These types of antennas are larger but suit capsules which fit the size restraints of 11 mm in diameter and 28 mm in length. If we were to consider downsizing the capsule, we would have to consider backscatter communication (explained in Backscatter Communication below) or other modern communication solutions.
Knowing all this, we chose the following for the capsule RF transmitter and receiver components: CC115LRGPR RF transmitter by Texas Instruments, with dimensions of 4 mm x 4 mm.
Powering of the electrical system raises a few issues. First of all, the size of the capsule is limited to ensure its safety. Batteries used for these types of applications take up a lot of space. Silver oxide batteries are most commonly used in ingestible capsule applications, since they have less safety risks than standard Lithium-ion batteries (Steiger et al., 2019). Silver-oxide batteries usually offer 3 V at 80 mAh and are coin-shaped with 8 mm diameter and 2.5 mm thick (Kalantar-Zadeh et al., 2017).
Although there are promising up-and-coming techniques in the field of wireless capsules, such as backscatter communication, we wanted to incorporate a silver-oxide battery for our capsule, to ensure quicker implementation and vast product development. For future development, we discuss backscatter communication for powering and transmittance in the Backscatter communication section.
On the other hand, to overcome limitations regarding the battery life, we want to implement a sleep mode in the device to minimize power consumption in the non-target parts of the GI tract. This can be done by using the microcontroller’s timer to trigger a valve (Tsubaki et al., 2014). An additional save in power consumption would be done by implementing a magnetic reed switch within the capsule and its container. The magnet is included in the capsule container which, in a normally closed (NC) reed switch, keeps the electric circuit open. Once the capsule is removed from the container, its circuit becomes closed, allowing electric current to flow through the system.
When developing our product, we also found a very promising novel technique for capsule communication. As previously discussed, wireless communication consumes a lot of power --half of the capsule’s energy-- and requires large batteries which can take up a lot of space of the capsule --up to 40-50% of the capsule’s total space (Jameel et al., 2019; Vasisht et al., 2018).
As explained by Jameel et al. (2019), the backscatter system is composed of three parts: transmitter, receiver and carrier emitter. The carrier emitter radiates the signal, which the backscatter transmitter can then use to forward the message to the receiver. The backscatter device can harvest energy from the RF waves to activate the device. A power source can be combined to improve reliability. Finally, the modulated signal sent by the transmitter can be decoded by the receiver (Jameel et al., 2019).
Vasisht et al. (2018) created a new backscatter design, ReMix, that is aimed for deep tissue devices, such as ingestible capsules. Today, backscatter devices are mostly used directly under the skin, since deep-tissue backscatter has been challenging to develop due to human surface interference and signal deflection. ReMix is able to cover these issues and this or other similar technology would be an optimal choice for our capsule in the future.
Backscatter communication offers intriguing possibilities to improve our ingestible capsule. With this type of wireless power transmission, we can reduce the battery requirements and thus, free space and increase the time that the capsule is able to measure. As In-Body backscatter also enables the localization of the capsule, the implementation of this method would give us more accurate data, since we can locate the concentration of the metabolite to a certain part of the GI tract. The method is quite new and still under research, but in the future, this technology could open new doors and solve many problems related to communication and powering of our capsule.
For the purpose of our ingestible capsule, we looked into what materials can be used to manufacture the device. The materials needed for the capsule are the overall capsule material (where the components are placed), the cell carrier compartment (which is composed of the mold and semipermeable membrane) and possible cladding.
For the overall capsule material, important aspects to consider when deciding on the material are mechanical properties. The capsule needs to hold together for the time that it is being used. Transparency is also an important factor, since we need to be able to detect the fluorescence and send light from LEDs through a thin layer of the polymer. On the other hand, when entering a market with a new product, it would be favourable to use materials that are already used for similar applications. In our case, we would hope that the material is both durable and safe to use. In a paper, written by N.K. Mandsberg et al. (2020), a set of biocompatible materials has been discussed. Specifically for ingestible devices, biocompatible polymers mentioned were polymethyl methacrylate (PMMA), polycaprolactone (PCL), poly vinyl alcohol (PVA) and 3D printed high temperature resin with cellulose acetate. In another paper, the dissolution time of these polymers were discussed. The dissolution time for PCL when measured in vitro at pH 7.4 was 2-3 years (Wu Y. et al., 2020). For PVA the dissolution time, measured in the same conditions, is 3-14 days. They also discussed the tensile strengths of PMMA, PCL and PVA, with PMMA having the highest 37-45 MPa, PCL with 16 and PVA with 18-89. PMMA is the most suitable option due to its transparency, biocompatibility and high tensile strength. It is also a favourable option since it is highly processable and can be used for various fabrication methods (Mandsberg et al., 2020).
Arduino Prototype and Code
While researching the components and design of the capsule, we also conducted practical examinations. To get a more in-depth understanding of the capsule's electrical design, we built a prototype that consisted of the Arduino UNO -microcontroller board, LEDs, LDR and RF transceivers (Figures 3 and 4). First, we tested different LDRs to see how they work and what kind of data they actually produce. We tested the LDR functionality by developing an Arduino-program for a LED that fluctuated the brightness of it. Both the LDR and the LED were chosen so that they resemble the fluorescent light of our chosen GFP and a sensor that is capable of detecting the GFP fluorescence’s wavelength, respectively. We connected a RF transceiver to the system and then sent the concentration data to another RF transceiver. The complete prototype consisted of two parts. The first one would measure fluorescent light and send that data to the receiver. The second one creates fluorescent-mimicking light and also receives the data from the other system. The latter system can be attached to a computer and with Excel Data Streamer add-in, we can obtain the light intensity data (Figure 5).
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