Reproducibility of spectral-domain optical coherence tomography total retinal thickness measurements in mice

Abstract

Purpose: To test the reproducibility of spectral-domain optical coherence tomography (SD-OCT) total retinal thickness (TRT) measurements in mice.

Methods: C57Bl/6 mice were anesthetized, and three repeated volumetric images were acquired in both eyes with SD-OCT (250 A-scans × 250 frames × 1024 samplings), centered on the optic nerve head (ONH). The mice were repositioned between scans. TRT was automatically measured within a sampling band of retinal thickness with radii of 55 to 70 pixels, centered on the ONH by using custom segmentation software. The first volumetric image acquired in a given eye was used to register the remaining two SD-OCT images by manually aligning the en face images with respect to rotation and linear translation. Linear mixed-effects models were fitted to global and quadrant thicknesses, taking into account the clustering between eyes, to assess imprecision (measurement reproducibility).

Results: Twenty-six eyes of 13 adult mice (age 13 weeks) were imaged. The mean global TRT across all eyes was 298.21 μm, with a mouse heterogeneity standard deviation (SD) of 4.88 μm (coefficient of variation [CV] = 0.016), an eye SD of 3.32 μm (CV = 0.011), and a device-related imprecision SD of 2.33 μm (CV = 0.008). The superior quadrant had the thickest mean TRT measurement (310.38 μm) and the highest (worst) imprecision SD (3.13 μm; CV = 0.010), and the inferior quadrant had the thinnest mean TRT (291.55 μm). The quadrant with the lowest (best) imprecision SD was in the nasal one (2.06 μm; CV = 0.007).

Conclusions: Good reproducibility was observed for SD-OCT retinal thickness measurements in mice. SD-OCT may be useful for in vivo longitudinal studies in mice.

Figure 1.

Side (left) and top (right) views of the stage used for alignment of the mouse for SD-OCT imaging of the ONH. A coverslip was used to account for the steep curvature of the mouse cornea.

Figure 2.

Manual alignment of two SD-OCT en face images. (A) Reference SD-OCT en face image and (B) subsequent SD-OCT en face image. (C) Scan registered to the reference.

Figure 3.

SD-OCT en face image (left) with the black line indicating the location of the OCT B-scan (right). The B-scan demonstrates automated segmentation of the ILM (white line) and RPE (gray line) to obtain measurements. Vertical lines: disc margin.

Figure 4.

maps aligned to a single reference scan. (A, C, F) OCT en face images; (B, D, G) TRT thickness map with sampling region indicated as the region between red concentric circles. (E, H) TRT maps registered to the reference.