Wide-field multiphoton imaging with TRAFIX

Abstract

Optical approaches have broadened their impact in recent years with innovations in both wide-field and superresolution imaging, which now underpin biological and medical sciences. Whilst these advances have been remarkable, to date, the ongoing challenge in optical imaging is to penetrate deeper. TRAFIX is an innovative approach that combines temporal focusing illumination with single-pixel detection to obtain wide-field multiphoton images of fluorescent microscopic samples deep through scattering media without correction. It has been shown that it can image through biological samples such as rat brain or human colon tissue up to a depth of seven scattering mean-free-path lengths. Comparisons of TRAFIX with standard point-scanning two-photon microscopy show that the former can yield a five-fold higher signal-to-background ratio while significantly reducing photobleaching of the specimen. Here, we show the first preliminary demonstration of TRAFIX with three-photon excitation imaging dielectric beads. We discuss the advantages of the TRAFIX approach combined with compressive sensing for biomedicine.

Optimal compressive multiphoton imaging at depth using single-pixel detection

“…Recently, TF has come to the forefront with the realisation that spectrally dispersed light preserves spatial fidelity throughout scattering media due to the temporal pulse compression being supported only by the inphase, minimally scattered photons [10][11][12][13]. Wide-field TF MPM has been demonstrated as a novel option for correction-free imaging at a depth of up to seven scattering mean-free-path lengths with two-photon [10] excitation and may go further using three-photon [14] excitation modes. This advance (termed TRAFIX) was enabled with single-pixel detection, wherein structured patterns are sequentially projected onto the sample plane.…”

“…In this paper, we have demonstrated that the random Morlet basis, based on Gabor's minimisation of the uncertainty criteria, presents an elegant and optimal solution to compressive imaging in microscopy. Very recently, we have seen a flurry of demonstrations combining wide-field temporal focusing with single-pixel detection to achieve correction-free multiphoton imaging at im-proved depths [10][11][12]14]. Fortuitously, this geometry lends itself to CS, with prospects to increase acquisition speed and minimise photobleaching, which in itself TRAFIX minimises compared to standard point-scanning approaches.…”