Dynamic real-time subtraction of stray-light and background for multiphoton imaging. Academic Article uri icon


  • We introduce a new approach to reduce uncorrelated background signals from fluorescence imaging data, using real-time subtraction of background light. This approach takes advantage of the short fluorescence lifetime of most popular fluorescent activity reporters, and the low duty-cycle of ultrafast lasers. By synchronizing excitation and recording, laser-induced multiphoton fluorescence can be discriminated from background light levels with each laser pulse. We demonstrate the ability of our method to - in real-time - remove image artifacts that in a conventional imaging setup lead to clipping of the signal. In other words, our method enables imaging under conditions that in a conventional setup would yield corrupted data from which no accurate information can be extracted. This is advantageous in experimental setups requiring additional light sources for applications such as optogenetic stimulation.

published proceedings

  • Biomed Opt Express

altmetric score

  • 0.75

author list (cited authors)

  • Fernndez, A., Straw, A., Distel, M., Leitgeb, R., Baltuska, A., & Verhoef, A. J.

citation count

  • 1

complete list of authors

  • Fern├índez, A||Straw, A||Distel, M||Leitgeb, R||Baltuska, A||Verhoef, AJ

publication date

  • January 2021