Baseband for FP measurements

[xiaojun]

It says that Baseband is used for FP normalization, but not clear what is the Baseband actually and what does ‘Clear’ mean for the Baseband setting?

Should this number change with time? We saw this changed with time and the trend of FP seems not right with this setting. Should we use other settings for Baseband?

Thanks.

[harrison]

The Baseband is a normalisation factor for FP measurement, as you said. See Note S15 in the supplementary of our BioRxiv here:
https://www.biorxiv.org/content/10.1101/796516v1.supplementary-material
which discusses why and how it is calculated.

In this “Clear” just refers to the “Clear” filter being used to take the Baseband measurement.

The Raw baseband number should change with time (or rather, with cell density), as more cells leads to more scattered light.

Depending on the biological system you are looking at, you may observe fluorescence go down at the start of an experiment (even at constant OD) if you have inoculated cells from stationary phase, as it will take some time for FPs that were previously expressed to be diluted out as they start growing.

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