Introduction

Endoscopic microscopy is an emerging subcellular-resolution technology that enables in vivo histology [1]. Optical coherence microscopy (OCM) is a noninvasive biomedical imaging modality which combines the high transverse resolution of conventional confocal microscopy and cross-section imaging ability optical coherence tomography [2]. By completing one full-field OCM scan, en face images of different depth can be reconstructed from three-dimensional volume data, at much faster rate and efficiency than confocal microscopy [3]. To perform endoscopic OCM, common path OCT (CP-OCT) is a promising approach since the probe arm in CP-OCT can have arbitrary length and thus favorable for endoscopic application [4]. In addition CP-OCT eliminates need for chromatic dispersion and other phase distortions related to the mismatching between the reference and the probe arm, which is critical for obtaining high resolution images. In this work, we built and tested an all-fiber common-path Fourier-domain optical coherence microscopy (CP-FDOCM) system, with image resolution of 2μm × 9μm (transverse × axial) using a simple fiber probe.

Experiment

A schematic of the experimental set-up is shown in Fig. 1 where C is a 50/50 coupler and only one branch on the right side is used as the common path for signal and reference. An all-fiber probe P is used for 3D scanning and the reference signal comes from the Fresnel reflection at the right-angle cleaved fiber probe end. The probe is driven by a GPIB controllable 3D moving stage M, which performs B-scan (transverse) in Y direction and C-scan (transverse) in Z direction. The combined sample and reference signals are received by H, a high speed spectrometer (Ocean Optics HR-4000) with a CCD detector array with 3648 pixels and 699nm~891nm range. A-scan (axial) is in X direction and the A-scan signals are processed by the computer program based on FFT algorithm. An SLED (EXS8410-2413) with 840nm central wavelength and ~40nm spectral FWHM is used as the light source, which gives a theoretical in-air resolution of ~8μm.

Fig. 1. CP-FDOCM experimental setup

Results

Fig.2 (a) shows the A-scan data using a mirror as a sample, where we measured the in-air axial 3dB resolution of ~9μm. Fig. 2(b) shows an image of USAF target obtained using the CP-FDOCM system, with transverse scanning step of 1μm. The 6th element of the 7th group can be clearly indentified, which corresponds to a transverse resolution ~2μm. Then we scanned the epidermal cells of fresh onion sample, with both B-scan and C-scan steps as 2μm. Fig. 3(a) displays the 3D volume of 500μm × 500μm × 490μm, and Fig. 3(b)~(e) are en face images reconstructed from the volume data, with 10μm axial spacing between them. Fig. 3(f) shows a 200μm × 200μm area at the depth of 150μm beneath the surface, and we can clearly indentify the nucleus and cell walls. Compared to previous OCM work using bulk optics, our all-fiber CP-FDOCM system has a decent transverse and axial resolution while being much more compact and endoscopy applicable.

Fig. 3. (a) Scanned sample volume of onion epidermal cells (500μm × 500μm × 490μm); (b)~(e): En face images (500μm × 500μm) reconstructed from 3D data; (f) Nuclei and cell wall in 200μm × 200μm area at the depth of 150μm beneath the surface.

References

[1] R. Kiesslich, M. Goetz, M. Vieth, P. Galle and M. Neurath, "Technology Insight: confocal laser endoscopy for in vivo diagnosis of colorectal cancer,” Nat. Clin. Pract. Oncol. 4, 480-490 (2007).
[2] B. Bouma, G. Tearney, in Handbook of optical coherence tomography, (Marcel Dekker Inc., 2001).
[3] S. Huang, A. Aguirre, R. Huber, D. Adler and J. Fujimoto, “Swept source optical coherence microscopy using a Fourier domain mode-locked laser,” Opt. Express 15, 6210-6217 (2007).
[4] A. Tumlinson, J. Barton, B. Považay, H. Sattman, A. Unterhuber, R. Leitgeb and W. Drexler, “Endoscope-tip interferometer for ultrahigh resolution frequency domain optical coherence tomography in mouse colon,” Opt. Express 14, 1878-1887 (2006).