When to use
- Planning a multiplex fluorescence imaging experiment
- Evaluating spectral overlap between candidate fluorophores
- Selecting filter sets for a fluorescence microscope
- Teaching fluorescence spectroscopy fundamentals
ToolsConductScience tool
MicroscopyFree in-browser calculator
Fluorescence Spectra Viewer.
Interactive spectral overlay for 20 common fluorophores. Visualize excitation and emission curves, compare Stokes shifts, and identify crosstalk between channels.
PrivateData stays in your browser
LiveNo sign-up required
Validated2026-04-08
CitableMethods and citation included
Calculator
Results update in placeFluorophores (3/6)
Blue
Green
Yellow
Red
Far-Red
Near-IR
Excitation & Emission Spectra
Dashed = excitation, solid = emission
Fluorophore Summary
| Fluorophore | Ex Peak (nm) | Em Peak (nm) | Stokes Shift (nm) | Category |
|---|---|---|---|---|
| DAPI | 360 | 461 | 101 | Blue |
| GFP | 488 | 509 | 21 | Green |
| TRITC | 547 | 572 | 25 | Yellow |
Crosstalk Matrix
| DAPI | GFP | TRITC | |
|---|---|---|---|
| DAPI | --- | 29.6% | 0.1% |
| GFP | 69.3% | --- | 3.2% |
| TRITC | 0.0% | 3.2% | --- |
<5% (minimal) 5-15% (moderate)>15% (significant)
Do not use for
- As a substitute for measured spectra from FPbase or Chroma
- For absolute intensity or brightness comparisons
- For time-resolved or phosphorescence measurements
Gaussian approximation
Real spectra have asymmetric tails and shoulders. Use this tool for planning and quick comparison, not publication figures.
Environment matters
Spectra shift with pH, solvent polarity, and protein environment. GFP emission in cells differs from purified protein.
Brightness is not shown
This tool normalizes all spectra to 1.0. Actual brightness depends on extinction coefficient and quantum yield.
Filter sets shape signal
Your actual detected spectrum depends on excitation filter, dichroic mirror, and emission filter — not just the fluorophore.
1
Method
Spectra modeled as Gaussian curves parameterized by peak wavelength and FWHM. Crosstalk computed as fraction of emission energy within neighboring detection bands. 20 fluorophores spanning UV to near-IR.
2
Validated
Last validated 2026-04-08. Calculations are designed for planning and documentation support; verify procurement decisions against manufacturer specifications or institutional SOPs.
3
How to cite
How to Cite
ConductScience Fluorescence Spectra Viewer (v1.0). ConductScience, Inc. 2026. https://conductscience.com/tools/fluorescence-spectra-viewer
Lambert TJ. FPbase: a community-editable fluorescent protein database. Nat Methods. 2019;16(4):277-278.
Lichtman JW, Conchello JA. Fluorescence microscopy. Nat Methods. 2005;2(12):910-919.
Fluorescence Fundamentals
Fluorescence is the emission of light by a molecule that has absorbed light of a shorter wavelength.
Key concepts:
- Excitation: photon absorption promotes an electron to an excited state
- Emission: the electron relaxes back, emitting a longer-wavelength photon
- Stokes shift: the wavelength difference between excitation and emission peaks
- Quantum yield: fraction of absorbed photons re-emitted as fluorescence
- FWHM: full width at half maximum, describing spectral bandwidth
Multiplex Panel Design
Imaging multiple targets simultaneously requires careful fluorophore selection.
Best practices:
- Space emission peaks by at least 40-50 nm
- Match excitation to available laser lines (405, 488, 561, 640 nm)
- Minimize crosstalk using the overlap matrix
- Use spectral unmixing when overlap is unavoidable
- Common 4-color panel: DAPI + GFP/AF488 + TRITC/Cy3 + Cy5/AF647
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