Difference between revisions of "Team:UCL/Design"

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<h2>Design</h2>
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By talking about your design work on this page, there is one medal criterion that you can attempt to meet, and one award that you can apply for. If your team is going for a gold medal by building a functional prototype, you should tell us what you did on this page. If you are going for the <a href="https://2015.igem.org/Judging/Awards#SpecialPrizes">Applied Design award</a>, you should also complete this page and tell us what you did.  
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<h5>Gut-on-Chip</h5>
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<h4>Introduction</h4>
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<p>To demonstrate a functional prototype of our project, we decided to show our system working under real-world conditions simulated in the lab using a Gut-on-a-Chip design similar to the one described in:
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<br><a href="http://pubs.rsc.org/en/Content/ArticleLanding/2013/IB/c3ib40126j#!divAbstract">http://pubs.rsc.org/en/Content/ArticleLanding/2013/IB/c3ib40126j#!divAbstract</a>
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<br><a href="http://pubs.rsc.org/en/Content/ArticleLanding/2012/LC/c2lc40074j#!divAbstract">http://pubs.rsc.org/en/Content/ArticleLanding/2012/LC/c2lc40074j#!divAbstract</a>
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<br>
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<br>
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The idea is to model the rate at which our genetically engineered bacterial culture (E. Coli Nissle) grows and colonizes the gut, and to characterize its expression of 5-HTP, a serotonin precursor that acts as an anti-depressant, in the device. Dr. Chiang, from UCL’s very own Microfluidics Lab, has already drawn up the 3d design described in the attachment using SolidWorks.
 
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<img src="https://static.igem.org/mediawiki/2015/3/39/UCL_SW_design.png" alt="SolidWorks1" style="height:200px;">
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<h4>Note</h4>
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<img src="https://static.igem.org/mediawiki/2015/b/be/UCL_SW_design_2.png" alt="SolidWorks2"style="height:200px;">
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<p>In order to be considered for the <a href="https://2015.igem.org/Judging/Awards#SpecialPrizes">Best Applied Design award</a> and/or the <a href="https://2015.igem.org/Judging/Awards#Medals">functional prototype gold medal criterion</a>, you must fill out this page.</p>
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<p>Dr. Paul Sharp, who works with human intestinal epithelial cell models at Kings College London, has kindly agreed to collaborate with us on this project, give us the Caco-2 cells we need, and advice us on the best ways to culture them. We then decided to improve on the original Gut-on-a-Chip designed at Harvard University by making it a more realistic mimic of reality and more financially feasible.The new design doesn't require a porous membrane, and is inspired by Dr. Marco's (UCL Biochemical Engineering) bulging bioreactor. It has been designed under the guidance of Dr. Paul Sharp. In addition to replicating the peristaltic motion of the longitudinal muscles in the intestines like Harvard's design, this model will also replicate the motions created by circular muscles.
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<p>This is a prize for the team that has developed a synthetic biology product to solve a real world problem in the most elegant way. The students will have considered how well the product addresses the problem versus other potential solutions, how the product integrates or disrupts other products and processes, and how its lifecycle can more broadly impact our lives and environments in positive and negative ways.</p>
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<img src="https://static.igem.org/mediawiki/2015/9/9c/UCL_GOC_design1.png" alt="GoC Design1" style="height:320px;">
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If you are working on art and design as your main project, please join the art and design track. If you are integrating art and design into the core of your main project, please apply for the award by completing this page.
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<img src="https://static.igem.org/mediawiki/2015/b/be/UCL_GOC_Design2.png" alt="GoC Design2" style="height:320px;">
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The microfluidics device will be initially tested using Monkey Kidney Epithelial Cells, which are very similar to Intestinal Epithelial Cells (Caco-2). Experiments were carried to determine the optimum seeding cell density of the cells, and the time they need to adhere.
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<h4>Monkey Kidney Fibroblast Cell Culture:</h4>
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<img src="https://static.igem.org/mediawiki/2015/d/d6/08-25-2015_post_cd73_bead_trypsin_replate_0004.png" alt="Monkey kidney cells1"style="height:200px;">
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<img src="https://static.igem.org/mediawiki/2015/e/ea/08-25-2015_post_cd73_bead_trypsin_replate_0005.png" alt="Monkey kidney cells2"style="height:200px;">
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<img src="https://static.igem.org/mediawiki/2015/f/f5/08-25-2015_post_cd73_bead_trypsin_replate_0006.png" alt="Monkey kidney cells2"style="height:200px;">
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<img src="https://static.igem.org/mediawiki/2015/9/94/08-25-2015_post_cd73_bead_trypsin_replate_0007.png" alt="Monkey kidney cells2"style="height:200px;">
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<h4>Protocol for Determining Optimum Seeding Cell Density:</h4>
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Cells were pipetted into a 96 well plate with cell densities reducing by half in each following column (8 replicates)
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After 3 days, the cell confluency was checked under a microscope to determine the optimum level.
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<h4>Column: Cell Count</h4>
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<li>50000</li>
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<li>25000</li>
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<li>12500</li>
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<li>6250</li>
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<li>3125</li>
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<li>1563</li>
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<li>781</li>
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<li>391</li>
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<li>195</li>
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<li>98</li>
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<li>49</li>
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<li>Negative Control</li>
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</ol>
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<h4>Protocol for Determining Adherence Time:</h4>
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<br>
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Cells were pipetted into a 96 well plate at the optimum seeding density
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At intervals of 1 hour, the medium from 1 column was removed
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The cells were stained with DAPI, and cell counting was done under the microscope.
 
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Revision as of 02:06, 2 September 2015

Gut-on-Chip

Introduction

To demonstrate a functional prototype of our project, we decided to show our system working under real-world conditions simulated in the lab using a Gut-on-a-Chip design similar to the one described in:

http://pubs.rsc.org/en/Content/ArticleLanding/2013/IB/c3ib40126j#!divAbstract
http://pubs.rsc.org/en/Content/ArticleLanding/2012/LC/c2lc40074j#!divAbstract

The idea is to model the rate at which our genetically engineered bacterial culture (E. Coli Nissle) grows and colonizes the gut, and to characterize its expression of 5-HTP, a serotonin precursor that acts as an anti-depressant, in the device. Dr. Chiang, from UCL’s very own Microfluidics Lab, has already drawn up the 3d design described in the attachment using SolidWorks.


SolidWorks1 SolidWorks2

Dr. Paul Sharp, who works with human intestinal epithelial cell models at Kings College London, has kindly agreed to collaborate with us on this project, give us the Caco-2 cells we need, and advice us on the best ways to culture them. We then decided to improve on the original Gut-on-a-Chip designed at Harvard University by making it a more realistic mimic of reality and more financially feasible.The new design doesn't require a porous membrane, and is inspired by Dr. Marco's (UCL Biochemical Engineering) bulging bioreactor. It has been designed under the guidance of Dr. Paul Sharp. In addition to replicating the peristaltic motion of the longitudinal muscles in the intestines like Harvard's design, this model will also replicate the motions created by circular muscles.

GoC Design1 GoC Design2

The microfluidics device will be initially tested using Monkey Kidney Epithelial Cells, which are very similar to Intestinal Epithelial Cells (Caco-2). Experiments were carried to determine the optimum seeding cell density of the cells, and the time they need to adhere.

Monkey Kidney Fibroblast Cell Culture:


Monkey kidney cells1 Monkey kidney cells2 Monkey kidney cells2 Monkey kidney cells2

Protocol for Determining Optimum Seeding Cell Density:

Cells were pipetted into a 96 well plate with cell densities reducing by half in each following column (8 replicates) After 3 days, the cell confluency was checked under a microscope to determine the optimum level.

Column: Cell Count

  1. 50000
  2. 25000
  3. 12500
  4. 6250
  5. 3125
  6. 1563
  7. 781
  8. 391
  9. 195
  10. 98
  11. 49
  12. Negative Control

Protocol for Determining Adherence Time:


Cells were pipetted into a 96 well plate at the optimum seeding density At intervals of 1 hour, the medium from 1 column was removed The cells were stained with DAPI, and cell counting was done under the microscope.