Lasers in Surgery and Medicine 38:955–959 (2006) Spatial Refractive Index Measurement of Porcine Artery Using Differential Phase Optical Coherence Microscopy Jeehyun Kim, PhD, 1,4 * Digant P. Dave ´, PhD, 2,4 Christopher G. Rylander, PhD, 4 Junghwan Oh, PhD, 3,4 and Thomas E. Milner, PhD 4 1 Beckman Laser Institute, University of California, Irvine, California 2 Biomedical Engineering, University of Texas at Arlington, Texas 3 Division of Radiation Oncology, University of Texas, MD Anderson Cancer Center 4 Biomedical Engineering Department, University of Texas of Austin, Texas Background and Objectives: We describe a methodol- ogy to record spatial variation of refractive index of porcine renal artery using differential phase optical coherence microscopy (DP-OCM). Study Design/Materials and Methods: The DP-OCM provides quantitative measurement of thin specimen phase retardation and refractive index by measuring optical path- length changes on the order of a few nanometers and with a lateral resolution of 3 mm. The DP-OCM instrumentation is an all-fiber, dual-channel Michelson interferometer con- structed using a polarization maintaining (PM) fiber. Results: Two-dimensional en face dual-channel phase images are taken over a 150200 mm region on a microscopic slide, and the images are reconstructed by plotting a two-dimensional refractive index map as the OCM beam is moved across the sample. Conclusions: Because the DP-OCM can record transient changes in the optical path-length, the system may be used to record quantitative optical path-length alterations of tissue in response to various stimuli. A fiber-based DP- OCM may have the potential to substantially improve in vivo imaging of individual cells for a variety of clinical diagnostics, and monitoring applications. Lasers Surg. Med. 38:955–959, 2006. ß 2006 Wiley-Liss, Inc. Key words: phase sensitive optical coherence micro- scopy; refractive index; relative phase retardation INTRODUCTION Optical coherence tomography (OCT) has emerged as an important optical imaging modality in noninvasive medical diagnostics. OCT generates high-resolution images by utilizing the cross correlation between light backscattered from the sample and the reference reflector. Various technical approaches have been developed to improve OCT spatial resolution, imaging acquisition rate [1,2], and image quality [3,4]. Most of these techniques use fringe amplitude or a combination of fringe amplitude and polarization information in reflected light [5]. Magnitude of back-reflected light is dependent on the local refractive index gradients of the structural components comprising the test specimen. Polarization information basically reveals different phase retardation, caused by a tissue, between two orthogonal polarization states of light. Knowledge of local refractive index variations in biologi- cal tissue is not only important in tissue optics but also a major factor that determines fringe signal variation in conventional OCT. Average refractive index over relatively homogeneous tissue medium has been measured by Tearney et al. [6] and Knu ¨ ttel et al. [7]. The basic approach presented by Tearney et al. [6] was to adjust the reference mirror for maximum signal at a particular focus position within a tissue whereas Knu ¨ ttel et al. [7] adapted a moving fiber tip/collimating lens in the sample path in order to trace maximum signal variation. Two-dimensional bire- fringence mapping of porcine myocardium tissue was reported [6]. The DP-OCM is capable of measuring optical path-length changes on the order of a few nanometers (5 nm) [8] so that it can be applied to image two-dimensional local refractive index variation of thin specimens. This article presents a method to image refractive index distribution over a tissue specimen processed by standard histology sectioning methods and fixated on a microscope slide. MATERIALS AND METHODS DP-OCM Instrumentation and Signal Processing Details of the DP-OCM used for refractive index profiling can be found in references [8,9]. A simplified schematic diagram is shown in Figure 1. The DP-OCM system is essentially a dual-channel Michelson interferometer with two independent channels corresponding to orthogonal polarization modes of a polarization maintaining (PM) optical fiber. At the interferometer input, light emitted from a broadband source (l 0 ¼ 1.31 mm and Dl FWHM 60 nm) is coupled into a PM optical fiber-based Michelson interferometer. In the reference path, the fiber is spliced at 458 to the input PM fiber of a lithium niobate (LiNbO 3 ) Y- waveguide electro-optic phase modulator which varies the optical path length giving a stable 50 kHz carrier frequency. A rapid scanning delay line [3] is used to *Correspondence to: Jeehyun Kim, Beckman Laser Institute, University of California, Irvine, 1002 Health Sciences Rd. East, Irvine, CA 92612. E-mail: jeehk@uci.edu Accepted 11 August 2006 Published online 17 November 2006 in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/lsm.20407 ß 2006 Wiley-Liss, Inc.