Comprehensive volumetric confocal microscopy with adaptive focusing

Dongkyun Kang,Brett E Bouma,Hongki Yoo,Guillermo J Tearney,Priyanka Jillella

doi:10.1364/boe.2.001412

Abstract

Comprehensive microscopy of distal esophagus could greatly improve the screening and surveillance of esophageal diseases such as Barrett’s esophagus by providing histomorphologic information over the entire region at risk. Spectrally encoded confocal microscopy (SECM) is a high-speed reflectance confocal microscopy technology that can be configured to image the entire distal esophagus by helically scanning the beam using optics within a balloon-centering probe. It is challenging to image the human esophagus in vivo with balloon-based SECM, however, because patient motion and anatomic tissue surface irregularities decenter the optics, making it difficult to keep the focus at a predetermined location within the tissue as the beam is scanned. In this paper, we present a SECM probe equipped with an adaptive focusing mechanism that can compensate for tissue surface irregularity and dynamic focal variation. A tilted arrangement of the objective lens is employed in the SECM probe to provide feedback signals to an adaptive focusing mechanism. The tilted configuration also allows the probe to obtain reflectance confocal data from multiple depth levels, enabling the acquisition of three-dimensional volumetric data during a single scan of the probe. A tissue phantom with a surface area of 12.6 cm2 was imaged using the new SECM probe, and 8 large-area reflectance confocal microscopy images were acquired over the depth range of 56 μm in 20 minutes. Large-area SECM images of excised swine small intestine tissue were also acquired, enabling the visualization of villous architecture, epithelium, and lamina propria. The adaptive focusing mechanism was demonstrated to enable acquisition of in-focus images even when the probe was not centered and the tissue surface was irregular.

Highlights

The diagnosis of Barrett’s esophagus (BE), dysplasia, and intramucosal carcinoma remains an important clinical problem
The first-line imaging method used for examination of the esophagus, does not have the contrast or microscopic resolution required to reliably detect the morphologic changes associated with BE progression
The feedback signal S was converted to TTL pulses that drove the piezo-electric transducer (PZT) actuator: the absolute value determined the number of TTL pulses, and the sign the duty cycle (0.23 and 0.77 for positive and negative values of S, respectively)

Summary

Introduction

The diagnosis of Barrett’s esophagus (BE), dysplasia, and intramucosal carcinoma remains an important clinical problem. The diagnosis of BE progression currently relies on histopathologic examination of tissues obtained from random endoscopic biopsy This method only allows a very small fraction of the region at risk to be examined and often fails to represent the overall disease status [1]. In a clinical study of imaging esophageal biopsy samples, SECM has been demonstrated to visualize architectural and cellular features similar to those used for histologic diagnosis [21]. The spectrally encoded line was helically scanned across static cylindrical specimens with similar dimensions to the human distal esophagus. This device captured large-area confocal images at a fixed focal distance and needed to conduct multiple helical scans at different focal locations to acquire volumetric data [22]. We show that the new probe design enables the acquisition of optically-sectioned images at multiple imaging depth levels during a single helical scan of the probe, a capability that greatly simplifies the imaging procedure and provides three-dimensional confocal microscopy information

SECM probe

Adaptive focusing

Volumetric imaging

Results

Discussions