Adaptive optics in multiphoton microscopy: comparison of two, three and four photon fluorescence

Opt Express. 2015 Nov 30;23(24):31472-83. doi: 10.1364/OE.23.031472.


We demonstrate adaptive optics system based on nonlinear feedback from 3- and 4-photon fluorescence. The system is based on femtosecond pulses created by soliton self-frequency shift of a 1550-nm fiber-based femtosecond laser together with micro-electro-mechanical system (MEMS) phase spatial light modulator (SLM). We perturb the 1020-segment SLM using an orthogonal Walsh sequence basis set with a modified version of three-point phase shifting interferometry. We show the improvement after aberrations correction in 3-photon signal from fluorescent beads. In addition, we compare the improvement obtained in the same adaptive optical system for 2-, 3- and 4-photon fluorescence using dye pool. We show that signal improvement resulting from aberration correction grows exponentially as a function of the order of nonlinearity.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Equipment Design
  • Equipment Failure Analysis
  • Feedback
  • Fiber Optic Technology / instrumentation*
  • Image Enhancement / instrumentation*
  • Interferometry / instrumentation*
  • Lasers*
  • Lenses*
  • Microscopy, Fluorescence, Multiphoton / instrumentation*
  • Miniaturization
  • Reproducibility of Results
  • Sensitivity and Specificity