Difference between revisions of "Team:HZAU-China/InterLab"

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                 <div class="h1">Cell Measurement</div>
 
                 <div class="h1">Cell Measurement</div>
                 <p>Cell measurement was the most complicated experiment we’ve ever countered in the whole InterLab. It
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                 <p>Cell measurement was the most complicated experiment we've ever countered in the whole InterLab. It
 
                     nearly took us a whole week to complete and diagnose the data. Each step of the experiment was so
 
                     nearly took us a whole week to complete and diagnose the data. Each step of the experiment was so
 
                     crucial that it could impact the result to a detectable extent.
 
                     crucial that it could impact the result to a detectable extent.
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                     <img src="https://static.igem.org/mediawiki/2018/3/3d/T--HZAU-China--interlab06.png" width=100% alt="">
 
                     <img src="https://static.igem.org/mediawiki/2018/3/3d/T--HZAU-China--interlab06.png" width=100% alt="">
 
                 </div>
 
                 </div>
                 <p>After repeated experiments we choose the best data and upload to headquarters.</p>
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                 <p>After repeated experiments we chose the best data and upload them to headquarters.</p>
 
                 <div class="h2">Colony Forming Units per 0.1 OD600 E. coli cultures</div>
 
                 <div class="h2">Colony Forming Units per 0.1 OD600 E. coli cultures</div>
 
                 <p>The CFU experiment is another challenging mission for its large amount of work and complicated
 
                 <p>The CFU experiment is another challenging mission for its large amount of work and complicated

Revision as of 14:34, 15 October 2018

The HZAU-China team also participated in the InterLab project as we did in the past years. At first we read and analyzed the protocol provided by the headquarters carefully, then we checked out if we had all the essential material and equipment. After everything was well prepared, we formally carried out the experiment.

Calibration
OD600 Reference Point

To obtain the OD600 reference point of a particular plate reader is rather necessary since we can only obtain ABS from a plate reader which is different from OD gained from a spectrophotometer. The ABS data obtained from a plate reader is volume-dependent while the sample of a spectrophotometer are volume-fixed.

As the protocol instructed, we added 100μL LUDOX solution into wells A1, B1, C1, D1 while added 100μL ddH2O into wells A2, B2, C2, D2. Then, we measured OD600 of the samples in the plate reader. Data obtained are demonstrated in the sheet below:

LUDOX CL-X H2O
Replicate 1 0.057 0.036
Replicate 2 0.055 0.039
Replicate 3 0.056 0.038
Replicate 4 0.056 0.039
Arith. Mean 0.056 0.038
Corrected Abs600 0.018
Reference OD600 0.063
OD600/Abs600 3.500

The data showed that the OD600 reference point of our plate reader is 3.500.

Particle Standard Curve

The particle standard curve experiment aims to obtain the relationship curve between the ABS600 of the solution and the concentration of bacteria. To achieve this goal, we need to find a kind of material which resembles bacteria very much. The headquarters sent us with Silica Beads, a kind of little particle which met our needs.

We made up a series of solution of different but continuous magnitude. To make the experiment more precise, we also set up another three parallel groups. Figures obtained are shown below:

Fluorescence Standard Curve

The fluorescence standard curve experiment targets to obtain the relationship curve between the strength of fluorescence and the concentration of fluorescein which has similar excitation and emission properties to GFP.

We prepared a series of fluorescein solution of different but continuous magnitude. As every experiment required, we also set up 4 parallel groups. Then we measured the strength of the fluorescence of each sample. Graphs obtained are shown below:

Cell Measurement

Cell measurement was the most complicated experiment we've ever countered in the whole InterLab. It nearly took us a whole week to complete and diagnose the data. Each step of the experiment was so crucial that it could impact the result to a detectable extent.

However, we fixed up all the mistakes and completed the experiment satisfactorily. Data achieved are shown below:

After repeated experiments we chose the best data and upload them to headquarters.

Colony Forming Units per 0.1 OD600 E. coli cultures

The CFU experiment is another challenging mission for its large amount of work and complicated operations.

We diluted the bacteria solution to OD600 = 0.1, then made up a sort of magnitude as the protocol required.

After reconducting the experiment for three times, we successfully achieved the data required by headquarters and finished the InterLab satisfactorily. The CFU data are demonstrated below:

Replicate 1 Replicate 2 Replicate 3
Positive 1 0.084 0.0945 0.0875
Positive 2 0.098 0.084 0.0875
Negative 1 0.0945 0.084 0.0875
Negative 2 0.1015 0.098 0.1015
Positive 1
Replicate 1 TMTC 11 1
Replicate 2 206 18 6
Replicate 3 174 17 2
Positive 2
Replicate 1 TMTC 9 4
Replicate 2 228 25 8
Replicate 3 278 2 0
Negative 1
Replicate 1 TMTC 23 1
Replicate 2 370 28 10
Replicate 3 TMTC 11 1
Negative 2
Replicate 1 TMTC 89 1
Replicate 2 TMTC 16 0
Replicate 3 TMTC 48 4
Summary

We completed the work on punctual with satisfaction from Measurement Committee, which delighted us very much.

The only suggestion we want to submit is that, in the Cell Measurement Experiment, we don’t think it’s correct to put the sample at 0 hour on ice. Because even if you put it on ice, the bacteria will also definitely grow, which results in high systematical error in the experiment.

Experiments Improve
InterLab

Calibration

Cell Measurement

Summary

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