Two-Photon Absorption Calculator

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Two-photon absorption cross section in Goeppert-Mayer units (1 GM = 10^-50 cm^4 s photon^-1). Typical organic fluorophores: 1-1000 GM; engineered dyes can reach 10,000 GM or more.
GM
Continuous or time-averaged laser power delivered to the focus. For pulsed lasers, use the time-averaged power (peak power x duty cycle).
W
Excitation wavelength. For TPA the photon energy is half of the one-photon absorption peak, so TPA typically uses near-infrared light (700-1100 nm) to excite UV/visible transitions.
nm
Full width at half maximum of the focused Gaussian beam at the sample plane. Tighter focus gives higher intensity and more TPA events. Confocal and two-photon microscopes typically use 0.3-1 μm; flow cytometry setups use 10-100 μm.
μm
Duration of illumination. For a pulsed laser, this is the total measurement window multiplied by the duty cycle (pulse width x repetition rate).
s
Photon flux at focusSaturation likely
3.73 x 10^25 ph/(cm^2 s)

Peak photon flux at the beam centre (photons/cm^2/s)

Excitations per molecule1,462.3979
Peak intensity8.83 x 10^6 W/cm^2
Beam radius (1/e^2)8.49μm
Photon energy2.365x10^-19 J
1,462.3979 events/molecule
Negligible<0.001Low0.001-0.1Practical0.1-10High / Saturating10+

Expected 1462.40 excitations per molecule per exposure.

  • Each molecule is excited on average 1462.4 times per exposure, which may push the fluorophore toward saturation or photobleaching.
  • Your excitation wavelength is in the biological near-infrared window (700-1100 nm), where tissue scattering and autofluorescence are minimised.
  • TPA rate scales with the square of intensity: halving the beam area (tightening the focus by 1/sqrt(2)) quadruples the excitation rate.

Next stepThe 1/e^2 beam radius at focus is 8.49 μm. Reducing this by using a higher-NA objective or a shorter focal length lens will increase TPA efficiency quadratically.

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