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I am doing some calcium imaging to measure calcium levels in HeLa cells. To do this I have expressed cameleon calcium indicator (YC3.60) in the cells and I am using confocal microscope Fluoview for the live imaging. I am having trouble to fully understand some settings about the microscope. As indicated on the picture settings below, I am using 440nm wavelength. However since the argon laser does not emit 440nm but 458nm I do not understand what does this 440nm stands for? Also after performing the live cell imaging, I analyze the data ratiometically. Is is possible to tell from my confocal settings what would be the ratio of Channel2/Channel1 in nm? I suppose it is 530nm and 480 nm but I am not sure if this is correct. I have read different papers using confocal microscope but it is not very clear to me since this is my first time using it. I would appreciate it a lot if someone can help me with this. enter image description here

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  • $\begingroup$ Your best bet will be to ask the person who runs the facility - they will have an idea on the set up at your facility and what will work best for your needs. $\endgroup$
    – bob1
    Aug 15, 2023 at 21:02
  • $\begingroup$ I thought about that but for the moment I don't know yet someone I can ask there. $\endgroup$
    – Ke Keiss
    Aug 15, 2023 at 22:18

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