Difference between revisions of "Team:Oxford/Applied Design"

 
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     <h1>Applied Design</h1>
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<center><img class="img-responsive" width="200px;" src="https://static.igem.org/mediawiki/2017/7/7a/Applied_design_page.png"></center>
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     <h2>Introduction</h2>
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     iGEM encourages all teams to take their projects beyond the lab and to consider design using a holistic approach. The question “What is our real world problem?” has been a key consideration from the beginning and has guided our project throughout the summer.
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     To ensure we were putting our diagnostic device into context, we considered various aspects including safety, accessibility and socioeconomic factors in Latin America. Various design iterations were built upon over the course of the summer, influenced by discussions with experts from a range of disciplines - including blood coagulation, microfluidics and general diagnostic devices.
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    Having examined these design options more carefully, we propose a final design for our system which fulfills our criteria for a suitable diagnostic device.
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    <h2>Developing Our Design</h2>
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    <h3>Criteria for a suitable diagnostic device: considerations from the OpenPlant Forum</h3>
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    Our early design criteria was influenced by research into the challenges of designing new healthcare technologies in developing countries. Our research indicated that important considerations included the level of infrastructure present, the cost to the end-user and the amount of training required.
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    In order to gain an insight into various aspects of synthetic biology, members of our team attended the OpenPlant Forum in Cambridge, UK. Dr Tempest van Schaik gave a talk titled ’Designing Diagnostics’, using her expertise in the development of bench-to-bedside healthcare technologies and importance of the end-user experience. From her talk, 3 key points stood out to us:
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         <li>Understand your analyte</li>
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            <li>What is the context in which the kit will be understood?</li>
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            <li>How exactly does taking blood work? Will we transport the blood to a different place, or do a spot-test by the bedside?</li>
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        </ol>
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        <li>Understand the users of the kit</li>
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        <ol>
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            <li>How will they be given the kit?</li>
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            <li>Will they want to use it?</li>
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            <li>Do they want it?</li>
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        </ol>
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        <li>Understand the diagnosis procedure</li>
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        <ol>
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            <li>What problems are encountered from an end-user perspective?</li>
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            <li>Can we simulate a diagnostics procedure to discover an issues?</li>
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         </ol>
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     </ol>
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    <p>
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    Having discussed these findings as a team, we proposed a set of general criteria to guide our initial design brainstorming.
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    <h3>Our 4E’s Applied Design Framework</h3>
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    <p>
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    Applied design is an important component of most iGEM projects, and requires an integrated and holistic approach to ensure projects are considered from a ‘real-world’ perspective. Based on our findings from OpenPlant and research, the Oxford iGEM 2017 team came up with a framework for considering applied design - the 4 E’s (‘Effectiveness’, ‘Ease of use’, ‘Economics’ and ‘Environment &amp; Safety’).
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    This provides a structured method for applied design considerations, and we hope that this framework may prove useful for future iGEM teams.
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            <div class="four-e-box" style="background: rgb(190,15,52)">Effectiveness</div>
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            <ul>
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                <li>How long will it take to get a clear result?</li>
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                <li>How will we ensure the test it sensitive and specific?</li>
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                <li>How can we ensure it can be used at the bedside?</li>
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            </ul>
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        </div>
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        <div class="col-sm-3">
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            <div class="four-e-box" style="background: rgb(72,145,220)">Ease of Use</div>
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            <ul>
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                <li>What equipment would be necessary to use it?</li>
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                <li>How can we present the result clearly?</li>
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                <li>What level of training would be required for the end-user?</li>
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            </ul>
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        </div>
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        <div class="col-sm-3">
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            <div class="four-e-box" style="background: rgb(207,122,48)">Economics</div>
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            <ul>
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                <li>What materials will be used?</li>
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                <li>Will it be feasible to transport our device in large quantities?</li>
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                <li>How can we alter it to reduce the cost?</li>
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            </ul>
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        </div>
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        <div class="col-sm-3">
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            <div class="four-e-box" style="background: rgb(170,179,0)">Environment &amp; Safety</div>
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            <ul>
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                <li>Can we reduce risks associated with using GMOs in healthcare technologies?</li>
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                <li>How can we ensure our product is sustainable and environmentally friendly?</li>
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                <li>How will the end-user dispose of any used materials?</li>
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            </ul>
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        </div>
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    </div>
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    <h3>Cell-free Systems</h3>
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    <p>
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    One of the first things we realised when we first began to plan our kit was the current difficulties with using cells as part of our system; they would require a cold chain for transportation and maintenance of the culture once they had arrived in location and were being used for the kit. Whilst reading around synthetic biology in general we discovered a paper from Pardee at al. (2016), which described a method for freeze-drying cell lysate to then be used as a cell-free transcription/translation system. This was perfect, as there was no need for a cold chain, and the lysate could be produced cheaply and easily. At the open plant forum we received lots of ideas from talks by Jim Swartz and Keith Pardee, then we discussed our project with Keith Pardee. He gave us lots of practical advice on designing an optimal circuit for cell-free expression, and we decided to add pre-synthesised TetR to our kit rather than producing it in our kit, as we’d originally planned on doing.
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    </p>
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    <p>After further reading, including papers from Garamella et. al (2016), we further improved the theoretical design of our kit by discovering that we could use linear DNA from a PCR reaction for our kit, rather than plasmid DNA. This almost eliminates the risk of contaminating the environment, as bacteria will not take linear DNA up nearly as easily as plasmid.
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<span id="header-lightbox-title">APPLIED DESIGN</span>
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    </p>
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</div>
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    <p>The final DNA-based part of the kit would be:</p>
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    <ul>
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        <li>Cell lysate, which can be mass-produced</li>
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        <li>PCR product ofr our circuitry thtat produces the TEV protease,</li>
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        <li>The TetR with the specific cleavage sequence that will be produced separately and added to the reaction.</li>
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    </ul>
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    <p>These would all be freeze-dried in the well of our kit.</p>
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</br>
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<p>During our research into cell-free, we noted a lack of information regarding its use in the field. This led us to produce a report on cell-free technology, which you can find below.<p>
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    <div style="margin-top: 600px"></div>
  
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<p> <center> <a href="https://2017.igem.org/Team:Oxford/Cell_Free_Report"><img class="img-responsive" width="200px;" src="https://static.igem.org/mediawiki/2017/f/fc/Cell_free_report.png"></a></center></p>
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<h2 id="title">Introduction</h2>
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</br>
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<p>iGEM encourages all teams to take their projects beyond the lab and to take a holistic approach to design. The question “What is our real world problem?” has been a key consideration from the beginning, and has guided our project throughout the summer.</p><br>
 +
<p>To put our diagnostic device into context, we considered various aspects including safety, accessibility and socioeconomic factors in Latin America. Many design iterations were built upon over the course of the summer, influenced by discussions with experts from a range of disciplines, including blood coagulation, microfluidics and general diagnostic devices.</p><br>
 +
<p>Having thoroughly examined and evaluated various design options, we propose a final design for our system which fulfils our criteria for a suitable diagnostic device.</p><br>
  
−
<p> <center><img class="img-responsive" width="1000px;" src="https://static.igem.org/mediawiki/2017/3/31/T--oxford--applied--design--fig3.jpg"></a><h6>A diagram showing the production process for cell-free technology</h6></center></p>
+
<h2>What diagnostics for Chagas disease currently exist?</h2>
  
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    <h3>Initial Design Iterations</h3>
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<p>Diagnosis of Chagas disease is difficult, as the disease is mostly asymptomatic in the acute phase and for the majority of the chronic phase.</p><br>
−
    <p>
+
−
    Our starting point involved the detection of cruzipain in a sample of blood, however we needed to design an output system and a method for reading the output. By applying our criteria to a cell-free diagnostic system, we were able to propose some early design options. We discussed these as a group, compared the relative merits of each, and decided which would be most suitable to carry on with.
+
−
    </p>
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    <h4>Early Stage: Output Ideas</h4>
+
<p>We spoke to Dr Carol Lole Harris, who advised us on the practical and social considerations involved in the application of our diagnostic in Latin America, based on her experience working in Paraguay. We discussed the current tests available for Chagas and Carol emphasised that a spot test would be by far the best option for a multitude of reasons.</p><br>
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    <table class="table">
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            <thead>
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                <tr>
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                    <th></th>
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                    <th>Color Dye</th>
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                    <th>Clotting System</th>
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                </tr>
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            </thead>
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            <tr>
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                <th>Advantages</th>
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                <td>
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                    <ul>
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                        <li>Many current spot tests use colour as an indicator</li>
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                        <li>Clear presentation of the result</li>
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                    </ul>
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                </td>
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                <td>
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                    <ul>
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                        <li>Possible to design bacteria to produce a clotting inhibitor</li>
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                        <li>Can perform test on blood sample without requiring preparation</li>
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                    </ul>
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                </td>
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            </tr>
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            <tr>
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                <th>Disadvantages</th>
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                <td>
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                    <ul>
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                        <li>Difficult to distinguish against the colour of blood</li>
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                        <li>Can't isolate plasma easily</li>
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                    </ul>
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                </td>
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                <td>
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                    <ul>
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                        <li>May be difficult to visualise clotting</li>
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                        <li>Result may be too subjective</li>
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                    </ul>
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                </td>
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            </tr>
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        </tbody>
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    </table>
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    <p></p>
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    <p>
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    From our research we found that the ‘clot-buster’ drug streptokinase is naturally produced by bacteria. This inspired us to use the properties of blood to our advantage; our output could interfere with the blood clotting system to produce a result.
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    </p>
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    <p>
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    With a coloured dye output, visualisation of the result would require the plasma to be isolated; this would increase the complexity and cost of the test.
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    </p>
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    <p>
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    Furthermore, a key goal for our applied design was to ensure it could be used in a scenario with minimal training and limited resources. Findings from the National Congenital Chagas Program in Bolivia (2004-2009) showed that follow-up after diagnosis was a major difficulty in controlling the disease; as such, an immediate bedside test would be most ideal. Isolation of plasma would require resources which may not be available at the bedside/in all healthcare settings.
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    </p>
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    <p>
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    Having reviewed these two options, we came to the conclusion that a system based around blood clotting as an output would be most suitable.
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    </p>
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    <h4>Late Stage: Ideas to measure blood clotting output</h4>
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<p>Following further reading, we created a table of our findings to highlight the lack of a rapid and feasible diagnostic for congenital Chagas disease. </p><br>
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    <table class="table">
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                    <th></th>
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                    <th>Blood Collection Tube</th>
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                    <th>Microfluidics</th>
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                </tr>
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            </thead>
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            <tr>
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                <th>Advantages</th>
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                <td>
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                    <ul>
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                        <li>Equipment is easy to obtain</li>
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                        <li>Relatively cheap</li>
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                        <li>Would fit into current infrastructure</li>
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                    </ul>
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                </td>
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                <td>
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                    <ul>
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                        <li>Provides a quantitative measure of coagulation</li>
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                        <li>Decreases analysis time</li>
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                        <li>Can use a smaller volume of blood (fingerprick)</li>
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                    </ul>
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                </td>
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            </tr>
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        </tbody>
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    </table>
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    <p></p>
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    <ul>
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        <li>Blood Collection Tube: Our research showed that most blood collection tubes are lined with anticoagulation factors, in order to prevent blood from clotting. This inspired us to produce a collection tube lined with our freeze-dried cell-free system.</li>
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        <li>Microfluidics: Two papers published by Steckl et al. (Lab on a Chip 2014, Biomedical Microdevices 2017) inspired us to consider a new method to screen for blood coagulation. Steckkl and colleagues presented a cheap, point-of-care blood coagulation assay, which utilised microfluidics in a paper-based device.</li>
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    </ul>
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    <p></p>
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    <p>
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    Whilst simple, we decided that the blood collection tube method may not produce a clear result, which was an important consideration.
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    </p>
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    <h3>Blood-clotting System: Hirudin vs Bivalirudin</h3>
 
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    <p>
 
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    Hirudin is a 65-amino acid peptide produced by leeches, and is used widely in the medical field as an anticoagulant. It is mass produced and purified using recombinant technology; initially hirudin was proposed as the output of our DNA/OMV systems, as recombinant hirudin expression in E. coli was shown to be efficient from the literature.
 
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    </p>
 
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    <p>
 
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    Applying our 4E’s framework to our decision to use hirudin led us to explore cost-friendly alternatives. We came across bivalirudin, a congener of hirudin with a similar mechanism of action. However, importantly, bivalirudin is a smaller peptide at only 20-amino acids in length. As a result, our cost analysis showed that it would cheaper to synthesise bivalirudin chemically than to produce hirudin recombinantly. This cost-difference provides a significant advantage in ensuring maximal availability of our kit.
 
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    </p>
 
  
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     <h3>Integrating Our Ideas Into A Design</h3>
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<h6>Table 1: Main diagnostic methods currently used to diagnose Chagas disease</h6>
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     <p>
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<table class="table table-hover">
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We began by sketching ideas on paper and sharing our thoughts during a group meeting. Following advice from Dr Tempest van Schaik, we produced a simple cardboard model of our design to view it from the end-user’s perspective. This proved very helpful - our design originally included positive control and negative control indicators, but we realised that this created a bulky, complicated device requiring three blood samples, and would increase our costs.  
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     <thead>
 +
      <tr>
 +
        <th>Test</th>
 +
        <th>How it works</th>
 +
        <th>Benefits</th>
 +
        <th>Limitations</th>
 +
        <th>Suitable for newborns</th>
 +
      </tr>
 +
     </thead>
 +
    <tbody>
 +
      <tr>
 +
        <td>Whole parasite microscopy</td>
 +
        <td>
 +
        <ul>
 +
        <li>Preparation of Giesma blood smears</li>
 +
        <li>Visualised using light microscopy</li>
 +
        </ul>
 +
        </td>
 +
        <td>
 +
        <ul>
 +
        <li>Established method</li>
 +
        <li>Carried out by already trained professionals</li>
 +
        </ul>
 +
        </td>
 +
        <td>
 +
        <ul>
 +
        <li>Not suitable for the when there is little <em>T. cruzi</em> in the blood</li>
 +
        <li>Not always possible to differentiate between <em>T. cruzi</em> from <em>T. rangeli</em>, which does not cause disease in humans.</li>
 +
        </ul>
 +
        </td>
 +
        <td>Yes</td>
 +
      </tr>
 +
      <tr>
 +
    <td>Polymerase Chain Reaction (PCR)</td>
 +
    <td>
 +
    <ul>
 +
    <li>Molecular detection of <em>T. cruzi</em> DNA is performed using a combination of three real-time PCR assays.</li>
 +
    <li>Acceptable specimen types are EDTA blood, heart biopsy tissue or cerebrospinal fluid.</li>
 +
    </ul>
 +
    </td>
 +
    <td>
 +
    <ul>
 +
    <li>Allows high sensitivity in the acute phase</li>
 +
    <li>Allows the presence of <em>T.cruzi</em> to be accurately distinguished from <em>T. rangeli</em></li>
 +
    <li>Allows direct detection of infection and easy interpretation of results</li>
 +
    </ul>
 +
    </td>
 +
    <td>
 +
    <ul>
 +
    <li>High variation in accuracy and lack of international quality controls</li>
 +
    <li>High cost and complexity means it is not practical to use in a clinical practice</li>
 +
    <li>Further validation is needed to prove whether PCR is suitable to diagnose the  chronic phase of Chagas</li>
 +
    </ul>
 +
    </td>
 +
    <td>Yes</td>
 +
      </tr>
 +
      <tr>
 +
      <td>Serological tests</td>
 +
      <td>
 +
      <ul>
 +
      <li>Detection of antibodies against <em>T. cruzi</em></li>
 +
      <li>Includes techniques such as indirect fluorescent antibody (IFA) test, a commercial enzyme immunoassay (EIA) and immunochromatographic tests</li>
 +
      </ul>
 +
      </td>
 +
      <td>
 +
      <ul>
 +
      <li>Can be used for acute phase and chronic phase</li>
 +
      <li>High specificity and sensitivity</li>
 +
      <li>Commercialised and approved for use by WHO</li>
 +
      <li>Low-cost formats are available</li>
 +
      </ul>
 +
      </td>
 +
      <td>
 +
      <ul>
 +
      <li>Cross reactivity can occur with diseases, such as leishmaniasis and schistosomiasis </li>
 +
      <li>Performance of these tests is lower than reported by their manufacturer, especially against specific strains of <em>T. cruzi</em></li>
 +
      <li>Not suitable for immunocompromised patients and newborns</li>
 +
      </ul>
 +
      </td>
 +
      <td>No</td>
 +
  </tr>
 +
    </tbody>
 +
  </table>
 +
<br>
  
−
Ultimately, we determined that by keeping the device as streamlined and inexpensive as possible, healthcare professionals using the kit could simply repeat the test for any inconclusive results.</p>
+
<p>We also spoke to Professor Dias Borges Lalwani is professor of epidemiology at the Faculty of pharmaceutical sciences at UFAM, whose contact details were forwarded to us by the AQA Amazonas team. We were fortunate enough to be able to arrange a Skype meeting with Jaila, which helped us contextualise the problem of Chagas highlighting the difficulty of seeking diagnosis given non-specific early disease symptoms. </p><br>
  
−
<h2>Current Kit</h2>
+
<h2>Why congenital Chagas disease?</h2>
−
<p>Our current kit meets our criteria established from our 4E’s framework: it is effective, easy-to-use, economically viable and environmentally safe. A prototype version was designed using CAD software and 3D printed.</p>
+
<p>We spoke to Dr Alonso-Vega of the University of San Simón, Cochabamba who ran the National Congenital Chagas Disease Programme in Bolivia from 2004-2009, and is an expert in congenital Chagas. Dr Alonso-Vega’s Chagas Disease Program recommended the need for a new congenital diagnostic. She gave us specific guidance for implementation of our diagnostic device in Bolivia and confirmed the suitability of our congenital test in Bolivian hospitals, where most deliveries occur.</p><br>
  
−
[Annotated diagrams of current kit]
+
<p>We first contacted Professor Yves Carlier early on in our project to gain more information about the pathology of congenital Chagas disease. Professor Carlier is a researcher in infectious diseases and clinical immunology at the Université Libre de Bruxelles. He re-emphasised the need for a diagnostic for neonates, strengthening our resolve to focus on congenital Chagas disease. He also informed us about the benefits of diagnosing Chagas disease in neonates, such as that treatment with benznidazole cures around 100% of babies if given before one year of age but not later in life.</p><br>
  
 +
<p>We also learnt that Chagas treatment for under 15’s is free, giving us confidence that our diagnostic would be accessible and impactful in containing Chagas if provided in a hospital setting. Our discussion with her further emphasised the requirement for a rapid point of care test that would allow treatment of infected newborns to begin before they left the hospital.</p><br>
  
−
<h3>Using the kit<h3>
+
<h2>How did we develop our design?</h2>
 +
<p>Based on our findings from the OpenPlant conference, we established a set of criteria for our applied design - the 4 E’s (‘Effectiveness’, ‘Ease of use’, ‘Economics’ and ‘Environment & Safety’).</p><br>
  
−
<p> <center> <img class="img-responsive";" src="https://static.igem.org/mediawiki/2017/e/e9/Diagnostic_procedure_flowchart.png"></center></p>
+
<p>Prototyping involved extensive consideration of various design iterations. We progressed from paper to cardboard to 3D printing - at each stage of this process, we were able to iron out flaws in our design and to incorporate new features from our evaluations.
  
−
<h3>Review of our kit by healthcare professionals</h3>
+
<p><a href = "https://2017.igem.org/Team:Oxford/Applied_Design_Developing">Read more here.</a></p>
  
−
<p>We presented our prototype model to two healthcare professionals in order to re-evaluate our current kit. Recommendations gathered would be implemented into the future versions of our kit.</p>
 
  
−
<h4>Mrs Sarah Dragonetti (Registered Nurse)</h4>
+
<h2>What is our solution?</h2>
  
−
Findings:
+
<p>Our current kit meets our criteria established from our 4E’s framework: it is effective, easy-to-use, economically viable and environmentally safe. Furthermore, it incorporates many of the recommendations provided to us by the experts we contacted. Our solution is a rapid, point-of-care diagnostic for congenital Chagas disease which can easily be used by any healthcare professional.</p><br>
−
<ul>
+
−
<li> Flat, rectangular pipette hole fits well
+
−
<li> Timestrip would be a useful tool during busy periods
+
−
<li> Good size and good shape - feels intuitive
+
−
<li> A window would allow you to see whether the pipette was emptied, preventing someone from accidentally drawing blood back up
+
−
<li> A red case would make it difficult to see the blood through the window, so white or translucent casing would be better
+
−
<li> Unclear on actual device when to click together the two components
+
−
<li> Unclear whether pipette should stay in kit or be taken out (and when)
+
−
</ul>
+
  
−
<h4>Dr Ben Riley (General Practice)</h4>
+
<p>Following extensive design and development, we produced a 3D prototype version of our kit. We spoke to Tim Ring, the vice president of safe-tec sales and marketing who generously sent us MICROSAFE® pipette samples so we could test the compatibility of these with our kit design. These pipettes are advantageous for blood collection and allowed us to test our applied design more rigorously.</p><br>
  
−
Findings:
+
<p> Our 3D prototype design was refined by the healthcare professionals we contacted for evalutation. The insights from Sarah Dragonetti (Registered Nurse) and Dr. Ben Riley (General Practice doctor) were hugely useful in assessing and refining the practicality of our diagnostic kit, consequently improving our applied design.</p>
−
<ul>
+
−
<li> Kit is sealed so no worry about blood containment
+
−
<li> Ideal cost should be comparable to a one-use diabetes test strip (~£3)
+
−
<li> Size and shape is good for packaging and transport
+
−
<li> Distinctive shape will make it easy to identify
+
−
</ul>
+
−
These were very useful comments - they support some aspects of our current design, but also propose some changes which would further improve the end-user experience.
+
  
−
<h4>Overall recommendations:</h4>
+
<p><a href = "https://2017.igem.org/Team:Oxford/Applied_Design_Solution">Read more here.</a></p>
−
<ul>
+
−
<li> Make the case transparent: prevents pipette errors but is more cost-effective than a window
+
−
<li> Make the key instructions as clear as possible, and include these within the kit
+
−
<li> Evaluate the cost of the device (which we have presented below)
+
−
</ul>
+
  
−
<h2>Integration of our kit into society</h2>
 
  
−
We consider integration of our device into existing healthcare systems and current infrastructure a key challenge, and therefore a fundamental aspect of our applied design considerations.
 
  
−
<h3>Dialogue with Juan Solano and Alfons Van Woerkom of The Global Fund</h3>
+
<h2>How will we implement our design?</h2>
  
−
We contacted Juan Solano and Alfons Van Woerkom of The Global Fund, an international financing organisation which has worked to fund several large-scale projects in the fight against HIV/AIDS, TB and malaria. They introduced us to the concept of market and financial analyses, in order to outline opportunities for a new diagnostic to come to market and to optimise its implementation. We reviewed a set of key points:
+
<p>We consider integration of our device into existing healthcare systems and current infrastructure a key challenge, and therefore a fundamental aspect of our applied design considerations.</p><br>
  
−
<ul>
+
<p>We received invaluable advice from David Sprent, an expert in International Supply Chain, and Juan Solano and Alfons Van Woerkom, representatives of the Global Fund who advised us on evaluating costs of manufacture, transport and taxation of a kit for end use by the Bolivian healthcare system. </p><br>  
−
<li> Existing diagnostics on the market
+
−
<li> Information on the size of the market
+
−
<li> Regulatory framework of the country
+
−
<li> Well documented details of the unit cost
+
−
<li> Well documented details of the phases of production
+
−
<li> Delivery aspects (materials, transaction costs, depreciation)
+
−
</ul>
+
  
−
This will allow an optimal pricing strategy to be determined at the level of the product and its users. Furthermore, it provides recommendations to potential investors regarding the product’s investment feasibility.
+
<p>We consulted with HeLEX, (Centre for Health, Law and Emerging Technologies) and with InSIS (Institute for Science, Innovation and Society). InSIS researches and informs key contemporary and emerging issues and processes of social, scientific, and technological change. The insights from both HeLEX and InSIS were highly useful in evaluating the ethical and social issues related to our project, and heavily influenced our applied design.</p></br>
  
−
  <h3> Cost </h3>
+
<p>Professor Keith Pardee has worked on cell-free technologies, and he informed us how cell-free systems could solve a lot of issues around safety and cost.</p><br>
−
    For costing of our kit we were aided by contact with David Sprent, an expert in International Supply Chain, and Juan Solano and Alfons Van Woerkom of the Global Fund. They helped us with costing, but also with the considerations that have to be taken into account when importing things into Latin America. We used Bolivia as a case study, and imagined what the situation would look like if the country were to adopt our kit wholesale, testing all 163,000 babies born every year. The kit would be manufactured in the UK and then transported to Bolivia, as according to <a href="https://2016.export.gov/industry/health/healthcareresourceguide/eg_main_092224.asp">Export.gov</a> as of 2014 there was no local production of pharmaceuticals.
+
−
    <h4> Materials </h4>
+
−
    <a href="#demo" class="btn btn-info" data-toggle="collapse">Table of Costs</a>
+
−
  <div id="demo" class="collapse">
+
−
    <table class="table">
+
−
        <tbody>
+
−
          <thead>
+
−
              <tr>
+
−
                  <th>Component</th>
+
−
                  <th>Cost Per Kit ($)</th>
+
−
                  <th>Source </th>
+
−
              </tr>
+
−
          </thead>
+
−
          <tr> <td> Bivalirudin </td> <td> 0.007 </td> <td> <a href="http://www.selleckchem.com/custom-peptide-synthesis.html">Cost (Selleckchem)</a> </br> (Modelling told us we needed 50uM) </td></tr>
+
−
          <tr> <td> Sodim Citrate </td> <td> 1.23^10-9 </td> <td><a href="https://www.researchgate.net/post/How_much_38_citrate_do_we_need_to_add_to_anticoagulate_blood_for_platelet_rich_plasma">Amount</a> </br>
+
−
            <a href="http://www.sigmaaldrich.com/catalog/product/aldrich/w302600?lang=en&region=GB">Cost (Sigma Aldrich) </a>
+
−
              </td></tr>
+
−
          <tr> <td> Calcium </td> <td> 0.00311 </td> <td><a href="http://onlinelibrary.wiley.com/doi/10.1046/j.1538-7836.2003.00075.x/full">Amount</a> </br>
+
−
            <a href="http://www.sigmaaldrich.com/catalog/product/aldrich/449709?lang=en&region=GB">Cost (Sigma Aldrich)</a>
+
−
              </td></tr>
+
−
          <tr> <td> Tissue Factor </td> <td> 8.78*10^-9 </td> <td><a href="http://onlinelibrary.wiley.com/doi/10.1046/j.1538-7836.2003.00075.x/full">Amount</a> </br>
+
−
            <a href="http://www.abcam.com/recombinant-human-tissue-factor-protein-ab119148.html">Cost (abcam)</a>
+
−
              </td></tr>
+
−
          <tr> <td> Capillary Tube </td> <td> 0.074 </td> <td> <a href="http://www.sigmaaldrich.com/catalog/product/aldrich/z611182?lang=en&region=GB">Cost (Sigma Aldrich)</a></td></tr>
+
−
          <tr> <td> Injection Molding of Kit </td> <td> 0.685 </td> <td> <a href="http://www.custompartnet.com/estimate/injection-molding/?units=1">Cost (CustomPartNet)</a></td></tr>
+
−
          <tr> <td> Cardboard Box (70x30x50mm) </td> <td> 0.1 </td> <td> <a href="https://www.abcpackaging.co.uk/">ABCPackaging</a></td></tr>
+
−
          <tr> <td> Microsafe Pipette </td> <td> 0.15 </td> <td> <a href="http://www.safe-tecinc.com/microsafe.htm">Cost (Safe-Tec)</a></td></tr>
+
−
          <tr> <td> Timestrip </td> <td> 0.11 </td> <td> <a href="http://timestrip.com/">Cost (Timestrip)</a></td></tr>
+
−
          <tr> <td> Printed Instructions </td> <td> 0.021 </td> <td> <a href="">?</a></td></tr>
+
−
          <tr> <td> TetR </td> <td> 8.25*10^-5 </td> <td> <a href="https://www.mybiosource.com/prods/Recombinant-Protein/Tetracycline-repressor-protein-class-B-from-transposon-Tn10/tetR/datasheet.php?products_id=1056262">Cost (MyBioSource)</a> </br> (Modelling told us we needed 100nM)</td></tr>
+
−
          <tr> <td> Cell Lysate for DNA Reaction </td> <td> 0.9 </td> <td> <a href="http://www.cell.com/cell/fulltext/S0092-8674(16)31246-6?_returnURL=http%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2FS0092867416312466%3Fshowall%3Dtrue">Pardee et al. (2016)</a></td></tr>
+
−
          <tr> <td> DNAse 1 Inhibitor </td> <td> 0.007 </td> <td> <a href="http://www.molcells.org/journal/view.html?year=2005&volume=19&number=1&spage=54">Choi et al. (2005) </br> Cost was assumed same as bivalirudin</a></td></tr>
+
−
          <tr> <td> <b> Total Cost </B> </td> <td> <b> 2.06 </b> </td></tr>
+
−
        </tbody>
+
−
    </table>
+
−
  </div>
+
  
−
          <h4> Manufacturing Cost </h4>
+
<p>Dr Piers Millett is a Senior Research Fellow at the Future of Humanity Institute, and he introduced us to the concept of platform technologies and how this could be applied to our project. This provides a good way of fast tracking future developments because the platform will already have met regulatory approval, therefore allowing the device to be more rapidly adapted for other diseases.</p><br>
−
          <p> This was hard to estimate, given the unknowns in our kit, but we assumed we would ask a third-party to assemble the kit and this would lead to costs of around <b>$0.25</b> per/kit. </p>
+
  
−
          <h4> Transportation Costs </h4>
+
<p><a href = "https://2017.igem.org/Team:Oxford/Applied_Design_Implementation">Read more here.</a></p>
−
          <p> We decided that with a minimum shelf life of around a year for our test it would be pertinent to send kits once a quarter to Bolivia, otherwise we risked them expiring before being used. With a 50x30x70mm box for our kit around 8,000 can fit on a europallet after taking into account further packaging for the pallet. This means we'd be sending 5 pallets/per quarter, and we estimated that this would cost around $12,000 per shipment to get from the factory in the UK to hospitals in Bolivia. This equates to around <b>$0.30</b> per kit. </p>  
+
  
−
          <h4> Taxes </h4>
+
<h2>References</h2>
−
          <p> Bolivia imposes a <a href"http://haiweb.org/wp-content/uploads/2015/08/Taxes-final-May2011a1.pdf"> <b>13%</b> </a> tax on pharmaceutical imports into the country.
+
  
−
          <h4> <b>Total Cost</b> </h4>
+
<p>Carlier, Y. and Truyens, C. 2017 Maternal-fetal transmission of Trypanosoma cruzi. Second Edition, American Trypanosomiasis Chagas Disease: One Hundred Years of Research: Second Edition. Second Edition. Elsevier Inc. doi: 10.1016/B978-0-12-801029-7.00024-1.</p>
−
          <p> Totalling up all these costs, and then adding 25%, as was suggested to us by those we contacted, brings the total cost of our kit to around <b>$3.90</b>, which is significantly less than the current other options on the market. </p>
+
  
−
<h3>Dialogue with HeLEX</h3>
+
<p>Cencig, S. et al. 2012 ‘Evaluation of benznidazole treatment combined with nifurtimox, posaconazole or AmBisome?? in mice infected with Trypanosoma cruzi strains’, International Journal of Antimicrobial Agents. Elsevier B.V., 40(6), pp. 527–532. doi: 10.1016/j.ijantimicag.2012.08.002.</p>
  
−
<p> Consultation throughout with relevant stakeholders, including the Centre for Health, Law and Emerging Technologies (HeLEX) and Piers Millet, has brought some important difficulties to our attention. Key issues raised from our dialogue include: </p>
+
</div>
−
</br>
+
</body>
  
−
<ul>
 
−
<li> Dual-use technology in synthetic biology
 
−
<li> Management of data gathered from our device
 
−
<li> Transnational boundaries and international collaboration
 
−
</ul>
 
  
−
</br>
 
−
<p>You can read more about social, economic and political factors affecting our project on our <a href="https://2017.igem.org/Team:Oxford/HP/Silver">Silver Human Practices page</a> </p>
 
−
</br>
 
−
<p>Whilst some of these issues (e.g. dual-use) may not immediately appear applicable to a diagnostic device, biosafety and biosecurity should be considered by any groups developing new technologies using synthetic biology. A component of our Education and Public Engagement activities therefore involved approaching some of these issues in order to foster a ‘culture of responsibility’ - <a href="https://2017.igem.org/Team:Oxford/Engagement">you can read more about our activities here</a>. </p>
 
−
</br>
 
−
<p>We discussed how optimal integration of our device partly requires established guidelines to fill in gaps which may exist in current regulation. Using Bolivia as a case-study, we have produced a policy brief which summarises some of these findings, and proposes a flowchart showing our proposed optimal diagnostics strategy for Chagas disease.</p>
 
  
−
</br>
 
−
 
−
<p> <center> <a href="https://2017.igem.org/Team:Oxford/Chagas_Public_Policy"><img class="img-responsive" width="200px;" src="https://static.igem.org/mediawiki/2017/9/98/T--oxford--chagas_disease--button.png"></a></center></p>
 
−
 
−
</br>
 
−
 
−
<p> <center><img class="img-responsive" width="500px;" src="https://static.igem.org/mediawiki/2017/e/e6/Diagnostics_flowchart.png"></a><h6>A flowchart showing the optimal diagnostic strategy for congenital chagas disease using a rapid protease detecting kit.</h6></center></p>
 
−
 
−
</br>
 
−
 
−
<p>One concern raised by our dialogue with HeLEX included the importance of public engagement in addressing the awareness of Chagas disease. A public health campaign rolled out in regions of Latin America with the implementation of our kit could circumvent future issues surrounding consent and knowledge of the risks associated with Chagas. Most importantly, this would need to be translated to local languages, including Spanish, to increase access of information to local stakeholders. To this end, we have produced a draft example of a public health poster which is concise and easy to read.</p>
 
−
</br>
 
−
 
−
<p> <center><img class="img-responsive" width="250px;" src="https://static.igem.org/mediawiki/2017/6/69/Oxford_igem_public_health.png"></a><h6>A draft public health poster for Chagas disease, translated into Spanish</h6></center></p>
 
−
 
−
<h2>Future Vision for our Kit</h2>
 
−
<p>As our kit is modular, it will be able to be easily and cheaply adapted to diagnose different diseases; the cost of changing the disease is then only the input block, not also the output block. Our vision for the future is that a streamlined manufacturing process can be established which allows a rapid development of new diagnostic modules as people characterise specific proteases which are biomarkers for disease.<p>
 
−
 
−
</br>
 
−
 
−
<p>References:
 
−
Portable, On demand biomolecular manufacturing, Pardee
 
−
The All E. coli TX-TL Toolbox 2.0: A platform for cell-free synthetic biology Garamella et al. </p>
 
−
       
 
 
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Latest revision as of 14:49, 15 December 2017


APPLIED DESIGN

Introduction

iGEM encourages all teams to take their projects beyond the lab and to take a holistic approach to design. The question “What is our real world problem?” has been a key consideration from the beginning, and has guided our project throughout the summer.


To put our diagnostic device into context, we considered various aspects including safety, accessibility and socioeconomic factors in Latin America. Many design iterations were built upon over the course of the summer, influenced by discussions with experts from a range of disciplines, including blood coagulation, microfluidics and general diagnostic devices.


Having thoroughly examined and evaluated various design options, we propose a final design for our system which fulfils our criteria for a suitable diagnostic device.


What diagnostics for Chagas disease currently exist?

Diagnosis of Chagas disease is difficult, as the disease is mostly asymptomatic in the acute phase and for the majority of the chronic phase.


We spoke to Dr Carol Lole Harris, who advised us on the practical and social considerations involved in the application of our diagnostic in Latin America, based on her experience working in Paraguay. We discussed the current tests available for Chagas and Carol emphasised that a spot test would be by far the best option for a multitude of reasons.


Following further reading, we created a table of our findings to highlight the lack of a rapid and feasible diagnostic for congenital Chagas disease.


Table 1: Main diagnostic methods currently used to diagnose Chagas disease
Test How it works Benefits Limitations Suitable for newborns
Whole parasite microscopy
  • Preparation of Giesma blood smears
  • Visualised using light microscopy
  • Established method
  • Carried out by already trained professionals
  • Not suitable for the when there is little T. cruzi in the blood
  • Not always possible to differentiate between T. cruzi from T. rangeli, which does not cause disease in humans.
 Yes
Polymerase Chain Reaction (PCR)
  • Molecular detection of T. cruzi DNA is performed using a combination of three real-time PCR assays.
  • Acceptable specimen types are EDTA blood, heart biopsy tissue or cerebrospinal fluid.
  • Allows high sensitivity in the acute phase
  • Allows the presence of T.cruzi to be accurately distinguished from T. rangeli
  • Allows direct detection of infection and easy interpretation of results
  • High variation in accuracy and lack of international quality controls
  • High cost and complexity means it is not practical to use in a clinical practice
  • Further validation is needed to prove whether PCR is suitable to diagnose the chronic phase of Chagas
 Yes
Serological tests
  • Detection of antibodies against T. cruzi
  • Includes techniques such as indirect fluorescent antibody (IFA) test, a commercial enzyme immunoassay (EIA) and immunochromatographic tests
  • Can be used for acute phase and chronic phase
  • High specificity and sensitivity
  • Commercialised and approved for use by WHO
  • Low-cost formats are available
  • Cross reactivity can occur with diseases, such as leishmaniasis and schistosomiasis
  • Performance of these tests is lower than reported by their manufacturer, especially against specific strains of T. cruzi
  • Not suitable for immunocompromised patients and newborns
 No

We also spoke to Professor Dias Borges Lalwani is professor of epidemiology at the Faculty of pharmaceutical sciences at UFAM, whose contact details were forwarded to us by the AQA Amazonas team. We were fortunate enough to be able to arrange a Skype meeting with Jaila, which helped us contextualise the problem of Chagas highlighting the difficulty of seeking diagnosis given non-specific early disease symptoms.


Why congenital Chagas disease?

We spoke to Dr Alonso-Vega of the University of San Simón, Cochabamba who ran the National Congenital Chagas Disease Programme in Bolivia from 2004-2009, and is an expert in congenital Chagas. Dr Alonso-Vega’s Chagas Disease Program recommended the need for a new congenital diagnostic. She gave us specific guidance for implementation of our diagnostic device in Bolivia and confirmed the suitability of our congenital test in Bolivian hospitals, where most deliveries occur.


We first contacted Professor Yves Carlier early on in our project to gain more information about the pathology of congenital Chagas disease. Professor Carlier is a researcher in infectious diseases and clinical immunology at the Université Libre de Bruxelles. He re-emphasised the need for a diagnostic for neonates, strengthening our resolve to focus on congenital Chagas disease. He also informed us about the benefits of diagnosing Chagas disease in neonates, such as that treatment with benznidazole cures around 100% of babies if given before one year of age but not later in life.


We also learnt that Chagas treatment for under 15’s is free, giving us confidence that our diagnostic would be accessible and impactful in containing Chagas if provided in a hospital setting. Our discussion with her further emphasised the requirement for a rapid point of care test that would allow treatment of infected newborns to begin before they left the hospital.


How did we develop our design?

Based on our findings from the OpenPlant conference, we established a set of criteria for our applied design - the 4 E’s (‘Effectiveness’, ‘Ease of use’, ‘Economics’ and ‘Environment & Safety’).


Prototyping involved extensive consideration of various design iterations. We progressed from paper to cardboard to 3D printing - at each stage of this process, we were able to iron out flaws in our design and to incorporate new features from our evaluations.

Read more here.

What is our solution?

Our current kit meets our criteria established from our 4E’s framework: it is effective, easy-to-use, economically viable and environmentally safe. Furthermore, it incorporates many of the recommendations provided to us by the experts we contacted. Our solution is a rapid, point-of-care diagnostic for congenital Chagas disease which can easily be used by any healthcare professional.


Following extensive design and development, we produced a 3D prototype version of our kit. We spoke to Tim Ring, the vice president of safe-tec sales and marketing who generously sent us MICROSAFE® pipette samples so we could test the compatibility of these with our kit design. These pipettes are advantageous for blood collection and allowed us to test our applied design more rigorously.


Our 3D prototype design was refined by the healthcare professionals we contacted for evalutation. The insights from Sarah Dragonetti (Registered Nurse) and Dr. Ben Riley (General Practice doctor) were hugely useful in assessing and refining the practicality of our diagnostic kit, consequently improving our applied design.

Read more here.

How will we implement our design?

We consider integration of our device into existing healthcare systems and current infrastructure a key challenge, and therefore a fundamental aspect of our applied design considerations.


We received invaluable advice from David Sprent, an expert in International Supply Chain, and Juan Solano and Alfons Van Woerkom, representatives of the Global Fund who advised us on evaluating costs of manufacture, transport and taxation of a kit for end use by the Bolivian healthcare system.


We consulted with HeLEX, (Centre for Health, Law and Emerging Technologies) and with InSIS (Institute for Science, Innovation and Society). InSIS researches and informs key contemporary and emerging issues and processes of social, scientific, and technological change. The insights from both HeLEX and InSIS were highly useful in evaluating the ethical and social issues related to our project, and heavily influenced our applied design.


Professor Keith Pardee has worked on cell-free technologies, and he informed us how cell-free systems could solve a lot of issues around safety and cost.


Dr Piers Millett is a Senior Research Fellow at the Future of Humanity Institute, and he introduced us to the concept of platform technologies and how this could be applied to our project. This provides a good way of fast tracking future developments because the platform will already have met regulatory approval, therefore allowing the device to be more rapidly adapted for other diseases.


Read more here.

References

Carlier, Y. and Truyens, C. 2017 Maternal-fetal transmission of Trypanosoma cruzi. Second Edition, American Trypanosomiasis Chagas Disease: One Hundred Years of Research: Second Edition. Second Edition. Elsevier Inc. doi: 10.1016/B978-0-12-801029-7.00024-1.

Cencig, S. et al. 2012 ‘Evaluation of benznidazole treatment combined with nifurtimox, posaconazole or AmBisome?? in mice infected with Trypanosoma cruzi strains’, International Journal of Antimicrobial Agents. Elsevier B.V., 40(6), pp. 527–532. doi: 10.1016/j.ijantimicag.2012.08.002.