Evaluation of photorefractive holographic imaging through turbid media

We are pursuing high spatial resolution confocal imaging of reflected NIR light as a diagnostic tool. To determine the efficacy of confocal imaging for in vivo pathology, it is critical to know the maximum thickness of scattering tissue through which biological signals can be detected. This limit depends on three factors: the fraction of signal photons generated in the focal region, the attenuation of the light to and from the focal region, and the background signal generated outside the focal region. Previous measurements of this limit used signals from a mirror,l which are much stronger than those expected from biological sources. Our previous Monte Carlo simulations* found that the greatest sources of contrast in reflection confocal imaging are changes in the index of refraction. For the index mismatches found in cells (-0.05),3 using a system NA of 0.4 we estimated these changes could be detected through 2-3 optical depths (ODs) of scattering. In this work, we have measured the maximum depth in ODs at which physiologic index mismatches can be detected. A confocal reflectometer was constructed (NA = 0.8) as shown in Fig. l a with an axial resolution of 4 pm. Two types of samples were investigated: 1) background was measured from homogenous scattering samples and 2) signal plus background was measured from scattering samples containing a planar index mismatch located at variable depths. Scattering was generated by 1-pm diameter latex microspheres. Scattering coefficients were calculated using Mie theory. The background was assessed by translating optically thick samples of scattering gelatin through the focus of the objective while recording the detected signal as function of position. The measured background scans from samples with three different scattering coefficients are plotted in Fig. 2a as a function of optical depth, where OD = (pJ (Depth.) An exponential fit to the background data indicates that each is decaying at eCoD. The initial amplitude of the background signal (B,) is directly proportional to the scattering coefficient (Fig. 2b). These data suggest that the background signal is produced by photons initially scattered outside the focal volume. Confocal signals from inhomogeneous scattering samples were measured by scanning a layered phantom with known optical properties (Fig. Ib). An index mismatch of 0.05 or 0.1 between the scattering gelatin and the scattering index fluid (IF) was used as a signal source at variable OD. The appropriate background was subtracted from each measure(b)