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OPTICS LETTERS / Vol. 35, No. 16 / August 15, 2010
Time-gated optical projection tomography Andrea Bassi,1,* Daniele Brida,1 Cosimo D’Andrea,1 Gianluca Valentini,1,2 Rinaldo Cubeddu,1,2 Sandro De Silvestri,1 and Giulio Cerullo1 1
Istituto di Fotonica e Nanotecnologie (IFN-CNR), Politecnico di Milano—Dipartimento di Fisica, Piazza Leonardo da Vinci 32, I-20133 Milano, Italy 2 Research Unit, Italian Institute of Technology (IIT), Piazza Leonardo da Vinci 32, I-20133 Milano, Italy *Corresponding author:
[email protected] Received May 5, 2010; revised June 30, 2010; accepted July 7, 2010; posted July 16, 2010 (Doc. ID 128022); published August 10, 2010 We present an imaging technique that combines optical projection tomography with ballistic imaging using ultrafast time gating. The method provides high-resolution reconstruction of scattering samples and is suitable for threedimensional (3D) imaging of biological models. © 2010 Optical Society of America OCIS codes: 170.6960, 170.6920.
Optical techniques able to visualize biological tissues in their three-dimensional (3D) organization are of great interest for the study of basic problems in life sciences [1]. High-resolution imaging techniques such as confocal or multiphoton microscopy are routinely applied, but their small field of view can make acquisition time too long for full tomography of samples on the millimeter scale. Other techniques have been developed to overcome this problem, including optical projection tomography (OPT) [2] and selective plane illumination microscopy (SPIM) [3]. Among these, OPT is particularly promising owing to its relative simplicity. Similar to x-ray computed tomography, OPT is based on the acquisition of a sequence of optical transmission images through the sample, which is rotated at several angles. The acquired projections are combined to image the tissue in 3D, typically using a filtered backprojection algorithm [4]. In addition to transmission, fluorescence from selectively labeled tissues can also be collected (fluorescence OPT) and can be used to obtain functional/molecular information. OPT was originally developed as an intensity-based technique, but it was also extended to fluorescence lifetime imaging [5]. Recently, much attention has been devoted to the study of biological models whose dimensions range from a few millimeters to 1 cm (the so-called mesoscopic scale) [6]. The applicability of OPT to the study of mesoscopic samples is strongly limited by scattering, and it is often necessary to apply chemical clearing techniques to reduce light diffusion. Here, a technique that extends the applicability of OPT to larger highly scattering media is proposed. Our approach, based on nonlinear time gating for the selection of the ballistic photons, is referred to as time-gated optical projection tomography (TGOPT). First proposed in the early 1990s, ballistic optical imaging through scattering media has since been implemented with different optical schemes, which include Kerr gate [7], nonlinear upconversion [8], and parametric amplification [9] (for a review, see [10]). It consists of selecting the photons that propagate straight through the tissue (ballistic photons) by gating only a short temporal window of the transmitted light using a suitable nonlinear optical process. To our knowledge, time gating has been applied only to projective imaging. In fact, optical tomography with early photons has been investigated using 0146-9592/10/162732-03$15.00/0
only electronic gating, based on the employment of intensified cameras [11], but these techniques are currently limited by the temporal duration of the intensifier response (typically larger than 200 ps) and are therefore only appropriate for the reconstruction of the optical properties of large diffusing samples, with submillimeter resolution. In this Letter, we demonstrate a full tomographic approach for the study of scattering media based on nonlinear upconversion. Furthermore, we test the applicability of the technique to the study of scattering samples ranging in the few-millimeter scale, suggesting that the method is applicable to 3D imaging of mesoscopic specimens, such as adult zebrafish. The experimental setup starts with an amplified Ti:sapphire laser with an 800 nm central wavelength, 50 fs pulse duration, 1 kHz repetition rate, and 1 mJ of energy. The laser beam is first spatially filtered by focusing in a hollow fiber in order to improve the beam quality, and then collimated to about a 5 mm beam waist. The polarization of a small portion of the beam (power
