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<img src="" alt="[protocols table]"> | <img src="" alt="[protocols table]"> | ||
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− | <font size="-2"><b> | + | <font size="-2"><b>Table 1: OD<sub>600</sub> measurement</b> </font> |
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<img src="" alt="[flourescence table]"> | <img src="" alt="[flourescence table]"> | ||
</div> | </div> | ||
+ | <font size="-2"><b>Table 2: Fluorescein fluorescence measurement with gain 48 & 70.</b> </font> | ||
</div> | </div> | ||
<br> | <br> | ||
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+ | <p>The task was to find the optimal plate reader settings for the cell measurement on Day 3 through a fluorescein dilution with PBS and a 485 excitation & 531 emission fluorescence measurement. We came up with the problem, that the same gain settings could not be used for both, fluorescein and cell measurement as in each case one of the measurement only displayed 50% usable outcomes. Additionally, only a gain of 48 or less displayed a complete set of fluorescein values. But the producing company of our plate reader only recommends a gain range for 50-150. So not even gain 48 displays “usable” values. We found the possibility for wider detection ranges in the product help documents, but our plate reader model Synergy MX from BioTek did not offer this option. Problems with the dilution by pipetting could be excluded.</p> | ||
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+ | <div class="row"> | ||
+ | <div class="col section-image border border-secondary"> | ||
+ | <img src="" alt="[Fluorescein standard curve]"> | ||
+ | </div> | ||
+ | <div class="col section-image border border-secondary"> | ||
+ | <img src="" alt="[Standard curve (log)]"> | ||
+ | </div> | ||
+ | </div> | ||
+ | |||
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+ | <font size="-2"><b>Diagram 1 & 2: This diagrams display the amount of fluorescence according to the fluorescein µM concentration in 100 µl LB media per well.</b> </font> | ||
+ | |||
+ | <p>As values bigger than 100,000 are labeled as OVERFLOW and don’t represent a usable number, the excel algorithm can’t display them in a usable curves.<p> | ||
+ | |||
+ | |||
<div class="section container"> | <div class="section container"> |
Revision as of 16:48, 31 October 2017
INTERLAB MEASUREMENT STUDY
Reliable and repeatable measurement is key to compare and analyse data from different labs all over the world. To support the effort to create detailed protocols to measure fluorescence and improve the possibility of comparing data, the iGEM Team NAWI_Graz 2017 decided to participate in the Fourth International InterLaboratory Measurement Study in synthetic biology. The goal is to establish a GFP measurement protocol based on engineering principles.
Our tasks were to follow exactly the iGEM Plate Reader Protocol, fill in the prepared excel sheets and send the results to the iGEM headquarter.
Results
Calibration Protocols
1. OD600 Reference point
Protocol Fluorescein Fluorescence Standard Curve
The task was to find the optimal plate reader settings for the cell measurement on Day 3 through a fluorescein dilution with PBS and a 485 excitation & 531 emission fluorescence measurement. We came up with the problem, that the same gain settings could not be used for both, fluorescein and cell measurement as in each case one of the measurement only displayed 50% usable outcomes. Additionally, only a gain of 48 or less displayed a complete set of fluorescein values. But the producing company of our plate reader only recommends a gain range for 50-150. So not even gain 48 displays “usable” values. We found the possibility for wider detection ranges in the product help documents, but our plate reader model Synergy MX from BioTek did not offer this option. Problems with the dilution by pipetting could be excluded.
As values bigger than 100,000 are labeled as OVERFLOW and don’t represent a usable number, the excel algorithm can’t display them in a usable curves.
Cell Measurement Protocol
- Day 1 OD600 Measurement
- Day 2 Transformation
- Day 3 Overnight culture
InterLab
Bronze Medal Criterion #4
Standard Tracks: Participate in the Interlab Measurement Study and/or improve the characterization of an existing BioBrick Part or Device and enter this information on that part's Main Page in the Registry. The part that you are characterizing must NOT be from a 2017 part number range.
For teams participating in the InterLab study, all work must be shown on this page.