InterLab
Connecting labs across the globe.
Introduction
As in all of the natural sciences, repeatability and comparability of experiments and data are essential to future development. The same holds true for synthetic biology, where it is especially important, as labs require research from other labs to build off of. However, it is not uncommon for different labs to report measurements in different units, resulting in an inconvenient variability between labs. As a result, our team has decided to participate in the Fifth International InterLab Measurement Study, which aims to reduce systematic variability between labs. Specifically, we will be aiming to reduce variability in fluorescence measurements by normalizing to absolute cell count or colony-forming units (CFUs) instead of using optical density (OD).
Plate Reader Details
- Instrument Brand and Model: Thermo Scientific Varioskan Flash 4.00.53
- Measurements: Supports both absorption and fluorescence
- Pathlength correction: Disabled
- Optics: Both top and bottom can be used
- Temperature: Room temperature (around 25 degrees Celsius, varies slightly each time)
- Bandpass width: 12 nm
- Excitation wavelength: 485 nm
- Emission wavelength: 520 nm
Results
1. Calibration
|
LUDOX CL-X |
H2O |
| Replicate 1 |
0.063189 |
0.043089 |
| Replicate 2 |
0.0633062 |
0.045393 |
| Replicate 3 |
0.0637365 |
0.043681 |
| Replicate 4 |
0.064268 |
0.044405 |
| Arith. Mean |
0.064 |
0.044 |
| Corrected Abs600 |
0.019 |
|
| Reference OD600 |
0.063 |
|
| OD600/Abs600 |
3.234 |
|
Table 1: Calibration results with our OD600/Abs600 conversion factor
According to the calibration test, our calculated OD600 to Abs600 conversion factor is 3.234.
2. Particle Standard Curves
Figure 1(a): Particle standard curve
Figure 1(b): Particle standard curve on a logarithmic scale
Despite the particle standard curve being a linear line, converting it into a log scale makes the graph non-linear.
3. Fluorescein Standard Curves
Figure 2(a): Fluorescein standard curve
Figure 2(b): Fluorescein standard curve on a logarithmic scale
Both non-logarithmic and log scale for the fluorescein standard curve appear to be linear.
4. Colony-Forming Units
[Unit: CFU/mL]
- BBa_I20270 Culture 1, Dilution Replicate 1: 4.24×10^7
- BBa_I20270 Culture 1, Dilution Replicate 2: 4.16×10^7
- BBa_I20270 Culture 1, Dilution Replicate 3: 4.4×10^7
- BBa_I20270 Culture 2, Dilution Replicate 1: 8.56×10^6
- BBa_I20270 Culture 2, Dilution Replicate 2: 6.96×10^6
- BBa_I20270 Culture 2, Dilution Replicate 3: 8.8×10^6
- BBa_R0040 Culture 1, Dilution Replicate 1: 2.64×10^7
- BBa_R0040 Culture 1, Dilution Replicate 2: 1.52×10^7
- BBa_R0040 Culture 1, Dilution Replicate 3: 2.88×10^7
- BBa_R0040 Culture 2, Dilution Replicate 1: 8.88×10^7
- BBa_R0040 Culture 2, Dilution Replicate 2: 2.88×10^7
- BBa_R0040 Culture 2, Dilution Replicate 3: 3.28×107
These are the values calculated from multiplying the number of colonies formed on the agar plate by its respective dilution factor.
Collaboration
We welcomed the SHSU_China iGEM team to use our laboratory and instruments to perform their InterLab study in return for borrowing their DNA Distribution Kit Plate. For more details regarding our collaboration process, please click here to advance to our collaborations page.
Figure 3: SHSU_China team members working on their InterLab at our laboratory
Figure 4: Our team member helping SHSU_China setup the plate reader software
Figure 5: Our team member assisting SHSU_China during the dilution series
Conclusion
We wish to thank iGEM for providing detailed protocols, materials, and this wonderful opportunity to participate in the InterLab. We have thoroughly enjoyed the InterLab and hope that our findings will contribute and corroborate those across different laboratories.
