1. Maximise rate.

A primary function of the beads is to maximise reaction rate per bead volume; halving the radius of a sphere doubles its surface area:volume ratio. Many, small, bacteria-embedded agarose beads (to a technical limit) are therefore optimal, as the average bacterium is closer to the surface of each bead and being ‘used’ more efficiently. Substrate molecules which follow Brownian motion are more likely to collide with and be broken down by ‘outer’ bacteria. Similarly, on average, product molecules have a shorter path length to the surface and are likely to diffuse out faster:

Assuming ρ (a coefficient of bacterial density), to be independent of r (distance from bead center) and R (bead radius), avg. bacterium-surface distance =



Oxford iGEM 2014

2. Protect bacteria

For this system and others of its type, it is highly valuable to maximise local substrate concentration to the bacteria within the viable range of toxicity, especially while the viable concentration range to the strain remains a limitation to the breakdown rate (directly or indirectly).

In our case, the diffusion-limiting polymer chosen was cellulose acetate (as its synthesis from cellulose is straightforward and safe) for which we modeled diffusion data for variable polymer thickness (see below). Acetylation stoichiometry or even polymer type entirely, polymer density and methods of bead coating are among many variables that can be further researched and optimised for desirable diffusion coefficients.

As bacteria need direct access to water, yet DCM is water-soluble only up to ~200mM, for limited water-solubility substrates, as part of future research, we propose suspending beads at the interface of a biphasic mixture of the two by exploiting differences in density. In such a system, the immediate substrate ‘reservoir’ is essentially maximised.

For the purposes of this project we opted to construct beads less dense than water, as the aqueous DCM concentration of the biphasic system is more reliable, and we had yet to establish the robustness of the diffusion-limiting system to external fluctuations in [DCM].

Step 1: Designing and visualising the biosensor

The replaceable cartridge

This cartridge is where the reporting bacteria would be held. The basic design is a block of agarose containing the bacteria, surrounded by a thin film, encased in a solid plastic shell to make it more user friendly. This cartridge is therefore very cheap and simple as it would have to be replaced every time the reporting bacteria culture became unusable.

The design shown here is an initial idea that is aesthetically pleasing.

The housing for the electronics

This housing is designed to contain the replaceable cartridge. The GFP sensing circuit will be contained in the top half of the box shown here to allow the circuit (specifically the light sensing diodes and the blue LEDs (both shown in the 3D images below)) to have maximum exposure to the reporting bacteria. The design also allows the cartridge to have maximum exposure to the solution that it is sensing.

Oxford iGEM 2014

Step 2: Constructing the components in 123D Design

To construct the complicated shape of the biosensor housing, we had to 3D print two halves of the biosensor individually so that we could have hollow sections. This is because 3D printers can’t easily construct anything that isn’t fully supported. This is the one major drawback of this type of rapid prototyping. The files of this construction are shown below.



Therefore, when we 3D printed the cartridge, we couldn’t go for the fancy designs that are shown above in SolidWorks. Therefore we had to opt for a much more linear design. The files of this construction are shown below.



These files were then transferred into Meshmixer . This planned out the actual printing of the components and performed certain useful features such as strength analysis to allow us to spot any large weaknesses before printing took place.

Step 3: Finished product

Pictures of the finished biosensor are shown below. The idea is that this biosensor can then be used as a very cheap sensor that can give a basic binary answer to the question ‘is there a safe level of chlorinated solvents present?’ This safe level will ideally be the 5ppm that is the maximum safe level for drinking water in the UK.

Oxford iGEM 2014