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Ravi S. Jonnal, Omer P. Kocaoglu, Qiang Wang, Sangyeol Lee, Donald T. Miller; Imaging of Cone Mosaic Dynamics using Phase-resolved Adaptive Optics Optical Coherence Tomography. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5874.
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Changes in the optical length of the human cone outer segment (OS)--both in response to visible stimuli and as a matter of its daily course of renewal and shedding--have recently been revealed using adaptive optics (AO) and flood illumination [1,2]. Ultra-high resolution optical coherence tomography (UHR-OCT) produces volumetric images of the retina, making it a natural choice for measuring these changes. Unfortunately, the demonstrated axial resolution of UHR-OCT is 3 µm --larger than the OS length changes of interest. Here, we demonstrate that AO-UHR-OCT can be combined with a phase retrieval technique in order to probe changes in the OS length much smaller than the axial resolution of the UHR-OCT system alone.
We used an AO-UHR-OCT system , incorporating a high-speed camera (125,000 lines/s; Sprint, Basler) and broadband SLD (840 nm; Broadlighter, Superlum), to acquire volumetric images of 0.5° square patches of retina, 1.5° from the foveal center, in three subjects. The camera was focused on the cone photoreceptors, and the AO system corrected ocular aberrations down to near the diffraction limit for a dilated 6 mm pupil. Each volumetric image was acquired within 180 ms, and series of 15 volumes were collected at a time. Custom software (MATLAB, Python/NumPy/SciPy) was developed to automatically correct intra- and inter-volume motion artifacts, and to automatically identify and track the cones, over the several seconds of acquisition time, as well as between measurements separated by hours. The software also retrieved phase information from reflective layers in the complex valued volumetric image. Phase values within individual cones were retrieved, and statistical analysis was performed on these values.
We found that the phase of light returning from the highly reflective surfaces of the connecting cilia and posterior tip was strongly correlated among the a-lines within individual cones, using ANOVA (p<<.01). We determined that the phase noise within cones was under 1 radian RMS, which suggests sensitivity sufficient to detect length changes as small as 47 nm.
We have demonstrated a method of detecting changes in the length of the OS significantly smaller than the axial resolution of UHR-OCT. This technique represents a uniquely sensitive tool for detecting and measuring physiological processes in the living cone photoreceptor.References1 Jonnal et al., Opt. Express 15(24), 2007.2 Jonnal et al., Opt. Express 18(5), 2010.3 Cense et al., Opt. Express 17(5), 2009.
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