SPECTROSCOPY GROUP 1 Using Raman Spectroscopy to Detect Malignant Changes in Tissues Introduction Accurate, rapid and non-invasive detection and diagnosis of malignant disease in tissues is an important goal of biomedical research. Optical methods, such as diffuse relectance, luorescence spectroscopy, and Raman spectroscopy, have all been investigated as ways to attain this goal. Diffuse relectance utilizes the absorption and scattering properties of tissues, particularly from cell nuclei and stroma. Changes in the scattering properties of tissues arise as the tissue becomes more dysplastic 1, 2 due to variations in hemoglobin content 3 and neovascularization. 4 Fluorescence spectroscopy is also inluenced by the changes in the optical properties of tissues and has been used to diagnose dysplasia. 5-8 However, there are a number of disadvantages to these techniques, including the need for extensive sample preparation or excision, as well as low sensitivity and speciicity rates. 6, 9 Many research groups have instead used Raman spectroscopy to detect and diagnose disease in vivo without the need for tissue removal or the addition of exogenous agents. Raman spectroscopy, a method based on Raman scattering, is a powerful technique that can be applied to many tissue sites. Raman spectroscopy is a molecular-speciic technique that probes the vibrational or rotational transitions in chemical bonds and provides detailed information about the biochemical composition of a sample. 10 The sensitivity of this technique is so high that a Raman spectrum is effectively a precise ingerprint of the biochemical makeup of the tissue. Application Note © 2011 Princeton Instruments, Inc. All rights reserved. A probe-based Raman spectroscopy system has been developed to non-invasively obtain Raman spectra in vivo for our research. The overarching goals of our group are to use Raman spectroscopy to successfully detect and diagnose abnormal tissues regardless of a patient’s age, race, body mass index (BMI), or medical history. Nearly identical systems have been set up to acquire Raman data to study a variety of malignancies, such as cervical dysplasia, changes indicative of preterm labor in the cervix, skin cancer, colon cancer, and breast tumor margins. After acquisition, Raman spectra are calibrated to account for day to day variations and processed to subtract background luorescence and smooth noise. Lastly, statistical analyses are performed to determine if Raman spectroscopy is capable of diagnosing malignant areas. Setup and Methods A schematic and picture of one experimental setup is shown in Figure 1. It consists of an EMVision iber optic probe connected to a 785 nm diode laser (from Process Instruments, Inc. or Innovative Photonics Solutions), a Kaiser Optical Systems imaging spectrograph (Holospec, f/1.8i-NIR) and a back-illuminated, deep-depletion, thermo-electrically cooled Princeton Instruments CCD camera (Pixis 256BR). These systems are all controlled with a laptop computer using software provided by Princeton Instruments (Winspec). In most experimental protocols, the iber optic probe delivers between 80 and 100 mW of light onto the tissue with an integration time between 2-5 seconds. During the measurements, all room lights and the computer monitor are turned off. A spectral resolution of 8 wavenumbers (cm-1) is achieved using these components. Elizabeth Vargis, M.S. and Anita Mahadevan-Jansen, Ph.D. Department of Biomedical Engineering Vanderbilt University