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Marianna Pavlyha, Ronald Silverman, Daniel Rohrbach, Harriet O Lloyd, Jonathan Mamou; Quantitative Scanning Acoustic Microscopy of the Cornea. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4335. doi: https://doi.org/.
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Recent studies comparing ultrasound and optical coherence tomography (OCT) measurements of corneal epithelium show systematic differences. In this study, we measured speed-of-sound and other acoustic parameters in the pig corneal stroma and epithelium using scanning acoustic microscopy (SAM) to evaluate speed-of-sound as a potential explanatory factor.
Intact pig eyes were shipped overnight on ice. The corneas were excised with a 1-2mm rim of sclera and placed in a 20% dextran in 0.9% saline solution for ~36 hours. Corneas were pressurized in a Barron’s anterior chamber to 10 mmHg and scanned with a 35 MHz ultrasound probe. Thickness values for 4 dehydrated and 2 control corneas were recorded. The central corneas were then excised with a 9.5 mm diameter Barron’s corneal punch. Corneal buttons were flash-frozen in Tissue Tek and cryosectioned to 5-μm. SAM was performed with a custom system using a 480 MHz probe. Ultrasound data were acquired over a 3x3 mm zone at a 2.5 GHz sample rate (12-bit) at 1-μm intervals. Subsequent analysis enabled determination and display of acoustic impedance, speed of sound, attenuation and bulk modulus.
Dehydrated and control corneas averaged 0.624±0.024 and 0.725±0.035 mm in thickness, respectively. SAM images of 5 zones from each of 3 sections of dehydrated corneas demonstrated significantly lower speed-of-sound in the epithelium, 1574±14 m/s, than in stroma, 1729±3 m/s. Mean impedance values for epithelium and stroma were 1.66 and 1.58 MRayl, respectively, and mean bulk moduli were 2.63 and 2.73 GPa, respectively.
In this study, we demonstrate that epithelium has a significantly slower speed of sound than the standard corneal value of 1640 m/s. This explains why OCT determinations of epithelial thickness have been reported to be significantly thinner than ultrasound determinations. Therefore, the standard corneal speed-of-sound value is inappropriate for accurate measurement of epithelial thickness. The ability to characterize and image impedance, speed-of-sound and bulk modulus at the 4-μm lateral resolution of the SAM could shed light on biomechanical changes affecting the cornea in disease (keratoconus) and therapy (crosslinking).
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
SAM image representing significantly slower speed of sound in epithelium compared to stroma
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