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Whirling-fiber endoscope allows two-photon imaging

May 2006
Breck Hitz

Two-photon imaging could be a useful medical technique if a suitable in vivo probe were available. Such a probe must be capable of rapidly scanning across a reasonably large area, and it must effectively deliver the excitation radiation and, equally, effectively collect the fluorescence signal. In addition, it must be flexible and small enough to be practical in vivo.

Recently, scientists from the department of engineering at the University of Washington in Seattle demonstrated a probe that meets these criteria. It consists of an ∋8-mm length of optical fiber mounted on a tubular piezoelectric (PZT) transducer whose outer surface is divided into four quadrants (Figure 1). By applying sinusoidal, out-of-phase voltages to opposite pairs of PZT electrodes, the scientists caused the fiber to whirl in a spiral pattern, scanning the emitted beam across the surface to be imaged. The entire device, together with a gradient-index lens, was housed in a 2.4-mm-diameter package at the end of an endoscope.

Figure 1.
Scanning was accomplished in the endoscope by a tubular piezoelectric transducer that whirled the short length of fiber in a circular spiral pattern (left). The fiber, transducer and a gradient-index (GRIN) lens were housed in a 2.4-mm-diameter package at the end of the endoscope (right).

Because the instantaneous position of the fiber lagged behind the voltage that was applied to the PZT electrodes, the image deciphered from the drive voltage was distorted. The magnitude of the lag depended on the instantaneous radial position of the fiber tip, and the scientists were able to insert a reliable correction factor to eliminate the distortion in real time.

Single-mode excitation radiation was delivered to the sample through the core, and multimode fluorescence was collected into both the core and the inner cladding of the same fiber. The double-clad fiber, from Fibercore Ltd. of Southampton, UK, had a core diameter of 3.6 μm and a 0.19 NA. The 90-μm-diameter inner cladding had a 0.23 NA. Because the double-clad fiber provided both a larger area and a better numerical aperture than a conventional single-mode fiber, it collected fluorescence with much better efficiency.

A home-built ultrafast Ti:sapphire laser provided the excitation signal in an experimental demonstration of the probe. To minimize pulse broadening caused by material dispersion in the fiber, the scientists applied a negative chirp to the pulses with a grating-based pulse stretcher before inserting them into the fiber. A dichroic mirror separated the outgoing excitation radiation from the returning fluorescence signal. The signal was then detected with a photomultiplier tube, digitized and analyzed to generate an image of the sample.

They used fluorescent beads, breast cancer cells stained with a fluorescent dye and live breast cancer cells targeted with fluorescein-labeled monoclonal antibodies as samples in an in vitro demonstration (Figure 2). The frame rate for these images was about 2.6 Hz, and each frame consisted of 512 rings with 521 pixels per ring.

Figure 2.
The endoscope was used to take images of fluorescent beads (A,B), stained breast cancer cells (C) and live breast cancer cells targeted with fluorescein-labeled monoclonal antibodies (D). The blurriness in the image of the 2.2-μm beads indicates that the endoscope is approaching its lateral resolution limit.

To develop their prototype into an effective medical tool, the scientists plan to redesign the optics to enhance delivery and collection of light at the distal end of the endoscope. They also intend to replace the current detector with one that is better suited for the fluorescence wavelength.

Optics Letters, April 15, 2006, pp. 1076-1078.

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