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@iGEM_Paris

Light Signalling in Gels

OPTIC MODEL

Introduction

Medusa aims at optically controlling bacteria immobilized in a gel by targetting them with two intersecting lasers. Two questions are important to address to achieve this goal:

How far can a laser go in a gel?

What resolution can the laser achieve?

We present in this page our study of the optical properties of commonly used culture gels. We show that it is possible to obtain a clean light signal even at gel depths greater than 10cm.

Gel Choice

The optical properties of a gel depend on the gel type, its concentration and the nutrient source.

Table 1: Gel Included in this Study

	Alginate	Agar	Gelrite
Key advantage	Solidifies at room temperature by adding calcium ion	Most ubiquitous microbiology gel	Low absorbance
Recommended concentration	1.75-4%	1-1.5%	0.15-0.25%
Melting-Gelling Point	NA	0.5%:85-20C 1%:90-35C	0.5%:65-15 1%:100-35C
Key disadvantage	Properties depend on calcium ion diffusion which is hard to control	High absorbance	Melts less well than agar
Concentration studied	1% with LB	1% with LB	1% & 0.5% with LB

The optical properties of a gel depend on the gel type, its concentration and the nutrient source.

It is best to use a low gel concentration to reduce absorbance. Conveniently some gels have a higher melting point than solidifying point. This hysteresis phenomenon means that the gel concentration can be lowered and still be kept at in a solid state at 37C by pre-cooling them (to 6-8C) prior to warm incubation.

Alginate gelling is induced by calcium ions which crosslink gelling polymers at room temperature. To get a homogenous gel, calcium ions cannot be mixed with an Alginate solution but must diffuse from an underlying calcium source (1). Since the optical properties of Alginate depend on the amount of crosslinking, they ultimately depend on calcium diffusion which is a difficult parameter to control. It is important to note that keeping the same diffusion conditions (time, temperature, calcium source concentration etc.) is necessary to maintain identical alginate optical properties between experiments. In addition to the gelling agent, the nutrient source also impacts light absorbance. Conveniently, LB shows less absorbance than M9 media at most visible wavelengths and was therefore added to our gels for our study.

We initially attempted to extract the intensity landscape in the gel from a simple picture. The strong diffusion of light from the main laser beam observed on these pictures suggested that an important scattering was occurring in the gel (sfig xx). Since the scattering observed on gel pictures was questioning the viability of Medusa, we further studied the light landscape with a different strategy. Briefly, we captured the light diffusion at various gels positions on the X-axis (X-axis represents gel depth) by scanning the light intensity along the Z-direction of the gel.

Lasers used in our study

your text

Data Fitting and Interpretation

The laser is affected within the gel by scattering because photons are redirected after colliding with gel particles. Scattering causes the laser to diffuse and creates a light halo surrounding the main beam. We found that the intensity of this light halo described a normal distribution on the Z-axis which we fitted a Gaussian equation (See table). The spread of this distribution represents the diameter of the laser beam. In contrast to what the gel pictures suggested, the laser diameter within the gel was actually close to its initial diameter, prior to entering the gel. The large light diffusion observed on gel pictures was probably caused by internal camera corrections which automatically balanced areas of high and low light intensity. In contrast, the low scattering that we recorded manually is very encouraging as it shows that a high targeting precision can be obtained within a gel which will give a high resolution to Medusa control.
In addition, to scattering, the laser beam is also affected by absorbance when photons energy is absorbed by gel particles and converted into heat. As expected, the laser intensity followed an absorbance induced logarithmic decay which we modeled by rearranging Beer-Lambert’s law with the equation defining absorbance:

1+sin(x)^{2}+3