Team:ACIBADEM ISTANBUL/InterLab

Table of Contents

1. Introduction
2. Results and Observation
3. Fluorescein Experiment
4. Cell Measurement
5. Conclusion

INTRODUCTION

Progress happens when data collected overtime accumulates and gives birth to new data or a product. However, to accumulate the data, variability in previous datas should be reduced to a minimum so that it can be declared that everyone reach the same results with same protocols and labs are able to reliably build upon others’ work.
As the one of the biggest interlaboratory studies, the goal of the iGEM InterLab study is to identify and correct the sources of systematic variability in synthetic biology measurements and has been successfully doing so for the past 4 years.
This year, the ınterlab studies aim to tackle the variability issues in fluorescence measurements. There are two given options to solve this problem normalizing to absolute cell count or normalizing the colony-forming units (CFUs) instead of OD.

Results and Observation

Table.1 A table of ludox results which have been used as reference.
Ludox
LUDOX CL-X was used as a reference to transform the absorbance data into a OD 600 value.
Particle Standard Curve
A suspension made with silica microsphere, which have similar optical properties with the cells used, was used to construct the particle standard curve.
Theoretically, as they have similar optical properties, the values obtained from this dilution series of the silica microspheres should be approximately the same as the values that should be obtained from cells. Thus the results were used to estimate the numbers of cells in certain absorbances.
This experiment was done according to the iGEM InterLab Plate Reader Protocol.
Fig 1. Particle standard curve generated by measuring the concentration of silica beads.
Fig 2. Particle standard curve generated by measuring the concentration of silica beads in log scale.

Fluorescein Experiment

Plate readers give out fluorescence values in arbitrary units that vary from one instrument to other. Therefore absolute fluorescence values cannot be directly compared from one instrument to another. This experiment was done in order to compare fluorescence output of test devices between teams.

Fig 3. Fluorescein Standard curve generated by measuring the concentration of fluorescein.
Fig 4.Fluorescein standard curve generated by measuring the concentration of fluorescein in log scale.

Cell Measurement

Fig 4.Fluorescein standard curve generated by measuring the concentration of fluorescein in log scale.
Fig 4.Fluorescein standard curve generated by measuring the concentration of fluorescein in log scale.

Conclusion

This study was a valuable experience for the team . It gave us valuable experience about fundemental lab techniques, teaching others, and problem solving issues.
During our collaboration with Saint Joseph, out team had to prepare the laboratory for people who were unexperienced in laboratory. Thus all reagents were pre-prepared the night before, the glassware autoclaved etc.
Also a safety lesson were given to them. During the lecture we were able to apply what we have learned from our own safety lecture. Problems encountered during the experiments had to be solved primarily by the team and that gave us an invaluable experience about problem solving skills.
The results came out as expected. We were able to see the linear relationship between florescein and concentration. However most probably due to pipetting errors during the silica microbeads protocol, the standard curve had slight waves.