Team:CU Boulder/Design
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The BioStake
The general design of the stake was inspired this
paper published in Biotechnology
Progress which describes a miniature bioreactor to detect
toxicity by using bioluminescent E. coli. Our design, shown in the
image, will holds cells in a gel at the top of the long
ground-insertion portion and will include homeostatic controls on the
top of the stake. The outer filtration and pump aspects are not shown
in the design above, but will be included to protect the cells from
harmful environmental influences.
Biological Design Considerations
Cells which contain logic gates and take part in
cell-to-cell signaling demand a very specific environment in order for
the system to work properly. Since the logic gates are intended to be
used as information storage as to whether a contaminant has passed and
are designed so monitoring does not need to be continuous, the cells
must be able to survive and remain active for several days inside the
device. Furthermore, the cell-to-cell singaling requires that the cells
are suspended in such a manner that AHL diffusion is efficient and that
they can induce one-another in an efficient manner. In order to support
both of these, we designed a special hydrogel scaffold made out of
crosslinked polyvinyl alcohol. PVA hydrogels are commonly used in
biomaterials based on its hyrdophilic nature which can attract moisture
and nutrients to keep the cells alive, its pore size which allows for
the diffusion of proteins and sugars while holding the cells in a
relatively fixed position, and its biodegradability, which is
attractive in the context of making an environmentally-conscious
product. Additionally, by developing a hydrogel with relatively small
pore size such as with PVA, it can act as a filter to keep out larger
competetive bacteria that may infiltrate the device as well as prevent
solid sedimentation from blocking cell-to-cell interaction and nutrient
diffusion.
In early September, PVA
scaffolds were made which contained
cells with the pBad inducible promoter in front of the RFP gene, and
experimental results support the hypothesis that the engineered E. coli
were able to live inside and respond to arabinose when soaked in the
solution. Further experimentation described below is still necessary to
confirm that this
In addition to designing the spatial arrangment of the cells, it's also
important to consider elements that will allow for their survival over
the time period which the stake is in place to collect data. If E. coli cells are
not kept in highly sterile conditions within a narrow temperature,
humidity, and nutrient concentration range, they will not sustain a
functional cell count for the system to work properly. Therefore, a
control system that can sense and automatically adjust these conditions
must be incorporated. The proposed plan is to use a solar-powered stake
so batteries do not have to be replaced, creating a more sustainable
device that can be used repeadetly. However, the incorporation of such
technology could potentially drive up the cost of the BioStake which
would not be favorable considering its intended use is for quick, easy,
and low-cost results available to anybody. Nonetheless, it seems to be
a
feasible solution to work with the cells that we have already developed.
Designing for Environmental Functionality
The types of locations in which the BioStake will
be placed, and how it tracks contamination at the sites is crucial to
the design of the stake itself. To create the best device, it should be
able
to intake an air or water sample that is consistent with the
bioavaiable concentrations of fracking contaminants, while excluding
harmful components from affecting the bacteria. The BioStake would
contain a small pump that would attract water or air through a filter
wraped around the base of the stake, and then small tubes would draw up
the fluid to the cell-containing gel via capillary action.
Creating a User-Friendly Device
As described on the motivation page, the market
has high demand for a biosensor that is easy to use and available to
the general public to detect fracking contaminants. Apart from the
design of the pump and homeostasis mechanism, many components of the
stake are designed specifically to make the BioStake inexpensive and
easy-to-use.
The physical structure of the stake is intended to be simple enough
that the main components could be manufactured in a plastic injection
mold. It's also designed for the hydrogel to be held in the compartment
between the top and the long ground-insertion portion so that the user
can know whether or not napthalene is present based on visual
inspection of the device to see whether the gel has changed colors. The
stake itself would be less than 6 inches in length and would weigh less
than a pound. Research still has to be done to balance being small
enough to be portable, and long enough for it to be able to collect
continuous fluid samples from air, water, or soil in the environments
around fracking sites.
Future Directions for Development
Further experimentation is required to determine
the
feasibility of using PVA to allow AHL diffusion in cell-to-cell
signaling, extensive cell
growth studies to see if the system can continue. If we had more time,
measurements would be taken to determine cell growth rates over time,
and then the hydrogel chemistry could be adjusted to control nutrient
diffusion appropriately.
Additionally, the
proposed method to maintain homeostasis within the device that would
allow the cells to survive over long periods of time may be tedious to
maintain and the aspect of the design that is most vulnerable to
failure. It may be beneficial to move the design from E. coli into a
bacterium that can survive in much harsher conditions, perhaps a
bacteria that already thrives in natural water sources.
Ideally, this product could be prototyped and put
into mass production. However, without enough information to specify
exact design criteria, its difficult to develop an entrepreneurial
plan, despite environmental agencies and oil companies expressing
interest in the product.