Three-Dimensional Quantitative Phase Imaging: Current and Future Perspectives Nicoleta M. Dragomir*, Xiao Ming Goh and Ann Roberts School of Physics, The University of Melbourne, Melbourne, Australia, +61 3 8344 5085 ABSTRACT Great effort has been made in the recent past to develop new non-destructive imaging modalities for both two and three dimensional objects, based on the phase properties of a specimen. Quantitative phase tomography (QPT) is a hybrid technique that has been proposed to provide three-dimensional (3D) refractive index (RI) profiling of irregular phase objects by combining transverse phase measurements with traditional tomographic reconstruction techniques. This profiling is accomplished through measurements of sets of projections which are ultimately related to the RI values of the object’s transverse cross-section. This is particularly useful for 3D refractive index determination of specimens where staining is not appropriate or for materials that cannot be stained and is essential to many applications in photonics and biotechnology. This article reviews recent developments in quantitative phase tomography as they are presently available and suggests future applications based on current research on the 3D RI. The enabling elements for 3D QPT in the context of four key areas are discussed: the effect of the refractive index of the surrounding matching fluid, spatial resolution, phase accuracy and optimal defocus. Recent progress and future perspectives related to each of these areas is presented with regard to various test objects of known optical properties. Keywords: Phase imaging, refractive index, tomography, three-dimensional microscopy. 1. INTRODUCTION Three-dimensional (3D) microscopic imaging has undergone a revolution in the past two decades. This is largely due to improved technology involving all current imaging modalities: confocal scanning microscopy 1 , multifocal multiphoton microscopy 2 , axial tomography 3,4 and optical tweezers 5 . There is a niche for 3D imaging of phase objects that has driven a remarkable renaissance in recent years 6-9 . The index of refraction is an important property for various specimens: biological 1,4,7,10-13 and non-biological 9,14-18 . The renewed interest in studying the 3D spatial distribution is a consequence of its importance in determining the optical properties of a range of specimens of importance to various industries including pharmaceutical, biotechnology, chemical and forensic analysis. While is well known that, as a result of its optical sectioning capability as well as due to the various fluorescent labels currently available 1,19 , confocal microscopy is capable of producing good resolution 3D images. However, issues related to low levels of intensity 20 , inability to resolve features of thick objects 19 due to limited frequency response still plagues its full potential for high resolution 3D imaging 3 . Furthermore, it can not be used for imaging of transparent specimens without using specific fluorescent labels that may negatively affect, in certain cases, live specimens. It has been demonstrated that the lack of 3D resolution in confocal volume renderings can be avoided through the application of axial tomography 3 . Depicted in the Fourier space this technique has the potential to measure almost all accessible spatial frequencies of a sample while rotating the sample under testing. Furthermore, when applied in conjunction with confocal microscopy to fluorescently labeled specimens, it has been demonstrated that only a small number of rotation steps are necessary 4 . However, this technique is computationally intensive as it deals with several volume data renderings. In this regard, it is thus apparent that true 3D transmission imaging requires a tomographic approach. Several tomographic schemes for observing various facets of microscopic transparent objects in the optical range have been proposed. They include optical diffraction tomography 21-23 , axial confocal fluorescence tomography 3,4 , digital holographic microscopy 24,25 , phase shifting interferometry 11,26 and phase imaging 9,27 . A review of these techniques can be found elsewhere 9 . It is worth noting that quantitative measurements on the RI can be found only in few of them 9,11,24 . This article presents a review of some of the technical capabilities of QPT. Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XV edited by Jose-Angel Conchello, Carol J. Cogswell, Tony Wilson, Thomas G. Brown Proc. of SPIE Vol. 6861, 686106, (2008) · 1605-7422/08/$18 · doi: 10.1117/12.762619 Proc. of SPIE Vol. 6861 686106-1 2008 SPIE Digital Library -- Subscriber Archive Copy