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Thank you everyone for the input on the spectral detector calibration.
We have two groups using the system. One for 7 colour OPAL staining and one for Teal/Venus and GFP/mRFP FRET sensors.
They will both redo the reference spectra in the new year for their new data. For the OPAL staining they had noticed some crosstalk
after the calibration (using the old reference spectra).
For the FRET work the reference spectra were collected with the black level set incorrectly. Each PMT has a different offset and some were fine and others were clipping the data. Then the spectra were collected correctly. Finally, they will have to collect new spectra with the recalibrated system.
I think we will take some time and look at things carefully and perhaps write up a technical note. The user will store the data with the appropriate reference spectra so they could be unmixed in the future and the data can be reproducible.
The Airy Scan discussion is interesting too. Maybe a new QUAREP-LiMi working group!
Sincerely,
Claire
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