Comparable fluorescence measurements
iGEM’s Interlaboratory study has been contributing to improvement of reproducibility and reliability of common experimental assessment methods. Labs worldwide are invited to participate in the eighth edition of this study, so we decided to provide our share to this greater cause. This year, the study aimed to expand the scope of the assessed measurements to multi-color calibrations and fluorescence development over time in a 96-well plate.
This preliminary experiment aimed at obtaining calibration lines for the three colors that are used in subsequent experiments, as well as for optical density. Fluorescein, Sulforhodamine-101, and Cascade Blue were used as calibrants for green, red, and blue fluorescence, respectively. In addition, silica microspheres were used to calibrate optical density (OD).
Unfortunately, a production issue affected the silica microspheres provided in the kit. Our team had already performed the experiments when this issue was elucidated, so we had to submit our data as is. This rendered the calibration of OD measurements impossible. Refer to the iGEM Technology website for more information on the implications of this.
Due to time limitations, the results were not analyzed or plotted. But the data files with the obtained measurements are provided below:
This experiment aimed to calibrate the fluorescence of six devices encoding for various fluorescent proteins (green, red and blue and combinations of two) to the three calibrant dyes. Additionally, the cell density was correlated to the OD calibrant. This was performed to assess the reproducibility of the newly designed three-color calibration protocol for the Interlab 2022 study.
Due to time limitations, the results were not analyzed or plotted. But the data files with the obtained measurements are provided below:
Within the frame of this experiment, the six tested devices encoded two fluorescent proteins in two transcriptional units, which differed in order in the different devices. With this, the experiment aimed to assess the reproducibility of fluorescence measurements and to investigate whether the order of transcriptional units influences their expression levels.
Due to time limitations, the results were not analyzed or plotted. But the data files with the obtained measurements are provided below:
This experiment aimed to assess the lab-to-lab reproducibility of dynamic fluorescence and OD measurements in a 96-well plate. Due to unavailability of a fluorescence plate reader in our laboratory, we had to perform the standard version of the experiment, in which the measurements were conducted at 0h and 6h.
Due to time limitations, the results were not analyzed or plotted. But the data files with the obtained measurements are provided below: