PHOEBE, a prototype scanning laser‐feedback microscope for imaging biological cells in aqueous media

Abstract

SUMMARY Based on the principle of laser‐feedback interferometry (LFI), a laser‐feedback microscope (LFM) has been constructed capable of providing an axial (z) resolution of a target surface topography of ∼ 1 nm and a lateral (x, y) resolution of ∼ 200 nm when used with a high‐numerical‐aperture oil‐immersion microscope objective. LFI is a form of interferometry in which a laser's intensity is modulated by light re‐entering the illuminating laser. Interfering with the light circulating in the laser resonant cavity, this back‐reflected light gives information about an object's position and reflectivity. Using a 1‐mW He–Ne (λ = 632·8 nm) laser, this microscope (PHOEBE) is capable of obtaining 256 × 256‐pixel images over fields from (10 μm × 10 μm) to (120 μm × 120 μm) in ∼ 30 s. An electromechanical feedback circuit holds the optical pathlength between the laser output mirror and a point on the scanned object constant; this allows two types of images (surface topography and surface reflectivity) to be obtained simultaneously. For biological cells, imaging can be accomplished using back‐reflected light originating from small refractive‐index changes (> 0·02) at cell membrane/water interfaces; alternatively, the optical pathlength through the cell interior can be measured point‐by‐point by growing or placing a cell suspension on a higher‐reflecting substrate (glass or a silicon wafer). Advantages of the laser‐feedback microscope in comparison to other confocal optical microscopes include: the simplicity of the single‐axis interferometric design; the confocal property of the laser‐feedback microscope (a virtual pinhole), which is achieved by the requirement that only light that re‐enters the laser meeting the stringent frequency, spatial (TEM00), and coherence requirements of the laser cavity resonator mode modulate the laser intensity; and the improved axial resolution, which is based on interferometric measurement of optical amplitude and phase rather than by use of a pinhole as in other types of confocal microscopes.

KTP crystal thickness distribution measurements based on laser feedback interferometry

“…The physical basis is the interference of the back-reflected field with the standing wave inside the laser resonant cavity. Hence, the optical feedback effect is also called self-mixing interference, and it has been studied hotly in the fields of displacement measurements [10,11], imaging vibration analyses [12], and microscopy [13].…”

Response of microchip solid-state laser to external frequency-shifted feedback and its applications

“…For micro-chip solid-state lasers, this value12 of the ratio reaches as high as ~10 6 . Consequently, these lasers can sense the extremely weak feedback power as low as 10 −12 , which permits broad application in fields such as tomography1213, microscopy14, interferometry15, vibrometry16, and velocimetry17. However, it is not yet clear to what amount of feedback power, the laser can be used for heterodyne phase measurement.…”