Difference between revisions of "Team:WashU StLouis/Interlab"

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<h1>Interlab</h1>  
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<!--<div class="action"><img  src="https://static.igem.org/mediawiki/2016/d/d9/T--WashU_StLouis--Action.png" style="width:100%;"></div>-->
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<h1 class="actionwords">Interlab</h1>  
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<h2 class="speechwords">We did the interlab study. &#9989; </h2>
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We did the interlab study. &#9989;
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<h5>WashU iGEM Interlab Study Results</h5>
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<p>This summer, in addition to our work on our iGEM project, we participated in the annual interlab study, a study whose goal is to compare measurements from different labs across the world. This year, the interlab study could be broken down to three basic parts: </p>
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<p>This summer, in addition to working on our iGEM project, we participated in the annual interlab study, which sought to compare measurements from different labs across the world. This year, the interlab study could be broken down to three basic tasks: </p>
 
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<li>Calibrating the OD600 by measuring with LUDOX</li>
 
<li>Calibrating the OD600 by measuring with LUDOX</li>
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<li>Measuring cell output fluorescence with 5 test devices</li>
 
<li>Measuring cell output fluorescence with 5 test devices</li>
 
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Below are our results for each part of the interlab study. At the end, we propose some improvements to the interlab study protocol.</p>
 
  
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<h5> Part 1 </h5>
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<p>OD600 correction using LUDOX
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<p>Below are our results for each part of the interlab study. At the end, we propose some improvements to the interlab study protocol.</p>
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**Reference OD600 = 0.01475<p>
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<h3> Part 1 </h3>
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<p>OD600 correction using LUDOX (Reference OD600 = 0.01475)<p>
  
 
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<h5> Part 2 </h5>
 
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<p>FITC Standard Curve</p>
 
  
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<p><img src="https://static.igem.org/mediawiki/2016/7/76/T--WashU_StLouis--Interlab_FITC_curve.png" /></p>
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<h3> Part 2 </h3>
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<p><img src="https://static.igem.org/mediawiki/2016/2/26/T--WashU_StLouis--Interlab1.png" style = "width:50vw; display:block; margin: auto auto;"></p>
  
 
<p>At this point in the interlab study, we encountered our first error. Our plate reader failed to measure fluorescence higher than 40,000 au. Therefore, we were unable to get results for highest five concentrations of FITC. See the end of this page to see our suggestions on avoiding this problem in the future. </p>
 
<p>At this point in the interlab study, we encountered our first error. Our plate reader failed to measure fluorescence higher than 40,000 au. Therefore, we were unable to get results for highest five concentrations of FITC. See the end of this page to see our suggestions on avoiding this problem in the future. </p>
  
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<h5> Part 3 </h5>
 
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<p>Cell fluorescence with 5 test plasmids</p>
 
  
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<p>Graph of Fluorescence values</p>
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<h3> Part 3 </h3>
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<p> <img src="https://static.igem.org/mediawiki/2016/0/0d/T--WashU_StLouis--interlab_fluorescence.png" /> </p>
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<p> <img src="https://static.igem.org/mediawiki/2016/8/83/T--WashU_StLouis--Interlab2.png" style = "width:50vw; display: block;margin: auto auto;""> </p>
  
 
<p>As seen in the graph above, fluorescence for all the devices increased as time went on, as expected. The negative control and device 3 both produced only small amounts of fluorescence.</p>
 
<p>As seen in the graph above, fluorescence for all the devices increased as time went on, as expected. The negative control and device 3 both produced only small amounts of fluorescence.</p>
  
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<h5>WHAT WE WOULD IMPROVE:</h5>
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<h3>What We Would Improve:</h3>
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<li>Further specifics should be given for measuring the FITC standard curve. For example, the protocol says to measure for GFP. Instead, a specific wavelength should be given.</li>
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<li>We are not sure if other teams had this issue, but we could not tell when the given FITC was properly resuspended. Due to a pipetting error, we lost our FITC and had to buy some from Thermo-Scientific. We believe that our store-bought FITC, which was not premeasured like the iGEM-supplied one, might have contributed to our wonky FITC curve. Having extra FITC would be helpful in case of an error.</li>
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<li>More parameters should be given for measuring the FITC standard curve. For example, the protocol says to measure for GFP. Measuring at different wavelengths could lead to different results, so wavelength should be specified for the study.</li>
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<li>We are not sure if other teams had this issue, but we could not tell when the given FITC was properly resuspended. Due to a pipetting error, we lost our FITC and had to use some from Thermo-Scientific. We believe that our store-bought FITC, which was not pre-measured like the iGEM-supplied one, might have contributed to our irregular FITC curve. Having extra FITC would be helpful in case of an error.</li>
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<li>Finally, we believe it is common practice to perform tests in triplicate, so we would recommend testing each colony in triplicate to get the best possible results. </li>
 
<li>Finally, we believe it is common practice to perform tests in triplicate, so we would recommend testing each colony in triplicate to get the best possible results. </li>
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Latest revision as of 19:43, 3 November 2016

MENU ▤

Interlab

We did the interlab study. ✅

This summer, in addition to working on our iGEM project, we participated in the annual interlab study, which sought to compare measurements from different labs across the world. This year, the interlab study could be broken down to three basic tasks:

  1. Calibrating the OD600 by measuring with LUDOX
  2. Finding a fluorescence standard curve with FITC
  3. Measuring cell output fluorescence with 5 test devices

Below are our results for each part of the interlab study. At the end, we propose some improvements to the interlab study protocol.

Part 1

OD600 correction using LUDOX (Reference OD600 = 0.01475)

Replicate 1 Replicate 2 Replicate 3 Replicate 4 Average Corrected Correlation Factor
LUDOX 0.0504 0.0553 0.0483 0.0511 0.051275 0.00765 1.928104575
Water 0.0433 0.0418 0.0472 0.0422 0.043625 N/A N/A

Part 2

At this point in the interlab study, we encountered our first error. Our plate reader failed to measure fluorescence higher than 40,000 au. Therefore, we were unable to get results for highest five concentrations of FITC. See the end of this page to see our suggestions on avoiding this problem in the future.

Part 3

As seen in the graph above, fluorescence for all the devices increased as time went on, as expected. The negative control and device 3 both produced only small amounts of fluorescence.

What We Would Improve:

  • More parameters should be given for measuring the FITC standard curve. For example, the protocol says to measure for GFP. Measuring at different wavelengths could lead to different results, so wavelength should be specified for the study.
  • We are not sure if other teams had this issue, but we could not tell when the given FITC was properly resuspended. Due to a pipetting error, we lost our FITC and had to use some from Thermo-Scientific. We believe that our store-bought FITC, which was not pre-measured like the iGEM-supplied one, might have contributed to our irregular FITC curve. Having extra FITC would be helpful in case of an error.
  • Finally, we believe it is common practice to perform tests in triplicate, so we would recommend testing each colony in triplicate to get the best possible results.