Corneal imaging with blue-light optical coherence microscopy

Abstract

Corneal imaging is important for the diagnostic and therapeutic evaluation of many eye diseases. Optical coherence tomography (OCT) is extensively used in ocular imaging due to its non-invasive and high-resolution volumetric imaging characteristics. Optical coherence microscopy (OCM) is a technical variation of OCT that can image the cornea with cellular resolution. Here, we demonstrate a blue-light OCM as a low-cost and easily reproducible system to visualize corneal cellular structures such as epithelial cells, endothelial cells, keratocytes, and collagen bundles within stromal lamellae. Our blue-light OCM system achieved an axial resolution of 12 µm in tissue over a 1.2 mm imaging depth, and a lateral resolution of 1.6 µm over a field of view of 750 µm × 750 µm.

Fig. 1.

Schematic diagram of the blue-light OCM system. SLED – super-luminescent light emitting diode; FC – 50:50 fiber coupler; PC – polarization controller; C1 - C3 – collimator; L1 - L4 – achromatic lenses; VFL – variable focus liquid lens; X, Y – galvanometer scanning mirrors; M – mirror; DM – dielectric mirror; VPHG – volume phase holographic grating; OL – objective lens; GPU – graphics processing unit.

Fig. 2.

(a) Ray-trace diagram from the blue-light OCM sample arm’s optical simulation in OpticStudio. (b) Spot diagrams with 750 µm × 750 µm FOV centered on the apex of the cornea model. Each spot diagram is encircled by an airy disk radius of 1.361 µm shown in black.

Fig. 3.

(a) SolidWorks mechanical layout of the sample arm. (b) Photograph of 3D printed mounts with optical components. (c) Sample (rabbit cornea) mounted on an artificial anterior chamber on a vertical stage shown in a red dashed box. C2 – collimator; L2 - L3 – achromatic lenses; VFL – variable focus liquid lens; DB – driver board for VFL; X, Y – galvanometer scanning mirrors; M – mirror; DM – dielectric mirror; OL – objective lens.

Fig. 4.

(a) Zemax simulation of the spectrometer with a zoomed inset of the rays reaching the camera emphasized with the purple box. (b) SolidWorks mechanical layout of the spectrometer. (b) Photograph of 3D printed mounts with optical components shown in the yellow dashed box. C3 – collimator; VPHG – volume phase holographic grating; L4 – focusing lens, f = 200 mm.

Fig. 5.

(a) Intensity falloff of up to an axial depth of 1.2 mm (b) PSF measurement of FWHM of ∼16 µm in air (∼12 µm in tissue). (c) En face view of a standard USAF 1951 resolution test target imaged with blue-light OCM. (d) A zoomed image of the yellow dashed box in (c) depicting the estimation of lateral resolution for group 9 and element 3 with 1.6 µm line spacing.

Fig. 6.

(a) Cross-sectional image of scotch tape to indicate image depth with the blue-light OCM system. Seven layers can be visualized with the aid of the variable focus liquid lens. (b) B-scan of a finger with visible sweat ducts. (c) Fungal filaments demonstrated on moldy bread. (d) E. coli bacteria in 1:10 glycerol solution (inset about 10× digitally zoomed in the orange dashed box).

Fig. 7.

En face images of rabbit corneal (a) endothelial cells, (b) collagen lamellae, and (c) keratocytes with 750 µm × 750 µm FOV. (Insets about 5× digitally zoomed).

Fig. 8.

Epithelium (a, d), endothelium (b, e) and keratocytes (c, f) in ex vivo rabbit cornea imaged with the blue-light OCM system (top row) and Heidelberg HRTII-RCM (bottom row).

Fig. 9.

(a-c) En face images of collagen lamellae in parallel stripes in ex vivo rabbit cornea. (d-f) Representative B-scans of each en face to emphasize the anterior and posterior sections of the fibers shown in green dashed boxes. The density of the fibers decreases as imaging depth increases.