System information

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3 Fluorescence
3.1 General Information
• Microplate Recommendations:
- Black plates are highly recommended.
- Clear plates can be used as a low-cost alternative if high signal intensity is expected.
- White plates are not recommended. It will increase noise by 20x to 50x.
• Samples with bubbles or high surface tension will affect fluorescence readings.
• Ensure the correct optical kit is used for the assay’s fluorophore.
• If the instrument was ordered with a luminescence module, a Microplate Sample Tray Cover
will have been installed. The Microplate Sample Tray Cover is not required for running in
fluorescence mode. For instructions on how to remove the Microplate Sample Tray Cover,
refer to Section 0 in the maintenance section.
• All values recorded are in raw RFUs (relative fluorescence units). To normalize, use the
Curve Fitting Data Analysis Software. See Appendix C for ordering information.
3.2 Optical Wavelengths and Commonly Used Fluorophores
Optical
Kits
Max.
Excitation
Wavelength
Emission
Wavelength
Typical Fluorophores
UV
365 nm 410 – 460 nm Hoechst dye, 4-MU
Blue
490 nm 510 – 570 nm EGFP, or AcGFP,
PicoGreen
®
, RiboGreen
®
,
Fluorescein, Quant-iT™
Protein, Quant-iT™
dsDNA
Green
525 nm 580 – 640 nm Rhodamine, Cy
®
3
Red
625 nm 660 – 720 nm Cy
®
5, Quant-iT™ RNA