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Lasers
| Laser | Typical Fluorophores | Dichroic Mirror Position | Notes | |
|---|---|---|---|---|
| 405 | DAPI, Hoechst | 405/532 | ||
| 488 | EGFP, FITC, Alexa 488 | 488/635 | ||
| 532 | Alexa 532, TRITC, Cy3 | 405/532 | This laser is not brilliant for exciting very red dyes like Alexa 594 and mCherry | |
| 635 | Cy5, Alexa 633, Alexa 647, Si Rhodamine dyes, DRAQ 5 | 488/635 |
Dichroic Mirror Position: There is a motorised dichroic mirror block that has to switch positions between 405/532 and 488/635 illumination paths, which will limit the speed with which green and red fluorophores (or blue and far red fluorophores) can be imaged. To optimise speed you should set up your multichannel imaging so the blue and red channels are acquired one after another and the green and far red channels are acquired one after another (e.g. blue/red/green/far red). That way the dichroic mirror only has to change position once between channels.