[vuthuhien97]

Hi Msilva and Harrison, I am facing a similar struggle as well. In my experiment, I tested various plasmids, ranging from green to red fluorescent, based on this paper with different emission/excitation settings. Surprisingly, despite adjusting the emission/excitation (Em/Ex) parameters to be far apart, I obtained green fluorescent signals even when the plasmid in the culture is expected to be red.
Also, I tested a non-fluorescent strain but still detected some fluorescence.
For my experiments, I used M9 medium and LED intensities ranging from 0.2 to 0.5. The plasmid used is a high copy number plasmid.

[vuthuhien97]

https://drive.google.com/file/d/1D-WdAEL0hgOEl0yujGJJCQH1Yba0w_DN/view?usp=share_link

• This reply was modified 1 year, 9 months ago by vuthuhien97.

[vuthuhien97]

Hi, Yes I mean the main LED. Yes, I tried plugging that reactor into other ports, and re-started the computer, but still. I only used fluorescent measurements and did not re-written the code.
The only changed is the LED intensity level was set at 0.5. However, I suppose that level is fine, am I right?
The link is the flashing light video of the reactor.
https://drive.google.com/file/d/1D-WdAEL0hgOEl0yujGJJCQH1Yba0w_DN/view?usp=share_link

• This reply was modified 1 year, 9 months ago by vuthuhien97.

[vuthuhien97]

No. I did not use the pumping in this case. I checked the troubleshooting guide, they did not mention the constantly flashing light.

[vuthuhien97]

Hello, we have increased the LED intensity level to 0.5 and it seems that we have detected a fluorescence signal. However, after running overnight, the reactor is no longer able to function; a light is constantly flashing. Even after I switched their connection, the control no longer detects that device. There are no problems with the data file. The file data looks ok same as the others. No error message. Do you have any ideas about what might be the cause of the problem? How could we fix it? Thanks so much.

[vuthuhien97]

Also, Do UI Data Plotting’s Normalised FP Emission graphs represent the fluorescence results? Could you please explain FP1 base and FP1 emit1 in the CSV data file? Which formula should I use to calculate fluorescence graph? It confuses me

[vuthuhien97]

Yes. I did, I used M9 media, I tried Chemostat as well. I always got the same results every time

[vuthuhien97]

Thanks so much for your suggestion. However, I still have trouble with the fluorescence signal. The data file of Chi.bio contains FP1 base and FP1 emit1. If I want to obtain the same fluorescent output as a Microplate reader. Which calculation must I perform? Thank you in advance!

[vuthuhien97]

Hi Harrison,
Thank you so much for your prompt reply
1, Yes, I used LB for both Chibio and the Microplate reader. Does this indicate that the Microplate reader detects the fluorescence signal with greater sensitivity than the Chibio device?

2, I started with a mixture of 100ul of overnight culture and 19,9ml LB fresh media. Did it achieve maximum fluorescence and growth???

I haven’t started with turbidostat growth or pump yet. I’ve only recently grown the bacteria in culture and measured the fluorescence signal.

Sorry, I don’t understand the inducible that you mention. How should I proceed? Could you please provide more information?
Thank you in advance!

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