Annular Beam Shaping in Multiphoton Microscopy to Reduce Out-of-Focus Background. (24th May 2017)

• Record Type: Journal Article
• Authors: Borglin, Johan; Wang, Danni; Durr, Nicholas J.; Hanstorp, Dag; Ben-Yakar, Adela; Ericson, Marica B.
• Other Names: Wong Kam-Sing Academic Editor.
• Is Part Of: International journal of spectroscopy. Volume 2017(2017)
• Publication Date: 2017-05-24
• Subjects: Spectrum analysis -- Periodicals; Spectrum Analysis; Spectrum analysis; Periodicals; Periodicals; 543.5
• DOI: 10.1155/2017/7560141
• Languages: English
• ISSNs: 1687-9449

Abstract

Abstract : Despite the inherent spatial confinement of multiphoton processes that arises from focusing through an objective, the maximum imaging depth in conventional multiphoton microscopy is ultimately limited by noise from out-of-focus fluorescence. This is particularly evident when imaging beyond shallow depths in highly scattering tissue as increased laser powers are necessary. The out-of-focus signal originates from multiphoton processes taking place primarily at shallow depths and deteriorates contrast and limits imaging depth. In this paper, annular laser beams are explored as a concept to reduce this background signal in multiphoton microscopy. The approach is theoretically verified by data from simulations and proof of principle is demonstrated on a custom-built experimental multiphoton microscopy platform. Annular laser beams were created by adopting wavefront control using a spatial light modulator and implemented for imaging tissue phantoms simulating turbid media and human skin ex vivo. The signal-to-background ratios were calculated and compared to images acquired with a traditional, filled-aperture Gaussian beam. Experiments in tissue phantom show an improvement in signal-to-background ratio of about 30% when using annular beam illumination in comparison to Gaussian illumination at specific depths. When laser power is not the limiting factor, this approach is expected to provide even greater benefits.