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How Are Acoustic-Optic Deflectors Used in Laser Scanning Microscopy?

Acoustic-optic deflectors play a central role in laser scanning microscopy and profilometry, offering numerous advantages over electro-mechanical scanners in terms of fast response time, high precision, and stable rasters.

Acoustic-optic deflectors based on single crystal TeO2 depend on anisotropic diffraction of light. This interaction requires more complex calculations than isotropic diffraction and must be undertaken with great care and accuracy.

Optical axis

An optical axis in a laser scanning microscopy system consists of components used to collect excitation laser light, fluorescence emission, and photon signals from specimens. These components include fluorescence filter sets and dichromatic mirrors as well as galvanometer-based raster scanning mirror systems with variable pinhole apertures to produce confocal images as well as photomultiplier tube detectors tuned specifically to different fluorescence wavelengths.

Laser scanning confocal microscope systems can be divided into two distinct groups. A single mirror system allows for straightforward x-y scanning with just one objective, while multiple objectives can be accommodated on one scan head – not possible with single mirror setups.

A typical commercial confocal scan head consists of a galvanometer-based raster scanning system with variable pinhole apertures for producing a confocal image, and photomultiplier tubes to capture any fluorescent emission from samples. An excitation laser beam passes through a fiber optic coupler and beam expander so as to fill up the entire rear objective aperture.

These deflectors are typically driven by an RF driver, with their diffraction efficiency proportional to the frequency of their generated acoustic signal. Their effectiveness may be restricted by material velocity; therefore, the usable angle range for their output beam may be restricted.

Acoustic-optic deflectors offer more drive frequencies than conventional modulators and can even be used with materials with slow sound wave diffraction coefficients, provided it can be maintained through acoustic control.

Acoustic diffraction is determined by two factors: frequency and travel time of an acoustic wave through material, which limits its amplitude; this limitation is particularly true for anisotropic diffraction which may prove challenging to achieve.

Manufacturers offer beam steering systems made up of piezo transducers and sound absorbers that utilize piezoelectric transducers for maximum diffraction efficiency, controlled via an analog control voltage analog RF driver to convert that voltage into an acoustic frequency; these devices are more stable than conventional modulators as the phase matching provided by piezo transducer/sound absorbers allows diffraction rates to match that of deflectors.

Scanning axis

Laser scanning microscopy employs multiple mirrors (typically two or three) to move a beam of laser across a sample and “descan” (transfer) it onto a pinhole and detector, producing high-resolution representative images of fixed samples. While the process can be slow, laser scanning microscopy provides accurate representations.

Confocal imaging is a versatile technique in confocal microscopy that often utilizes confocal geometry to mitigate light from other regions within a sample, particularly before and after its focal point. By doing so, depth resolution improves and, ideally, focal plane thickness can become very thin.

However, confocal point-scanning is limited by a finite velocity of sound in acousto-optic material that restricts its range of scan angles. An acousto-optic deflector may help increase this number.

An acousto-optic deflector is a modulator that sequentially changes the direction of a fixed laser beam based on an acoustic signal. Frequency control allows users to customize this deflection angle as sound velocity affects acoustic waves’ sound velocity; usually deflection angles tend to be small as optical wavelength is longer than its acoustic wavelength.

Acousto-optic deflectors come equipped with various acoustic frequencies and input aperture sizes, enabling a variety of scanning angles and use with various laser beams.

Laser scanning microscope systems often employ various types of scanners in addition to acousto-optic deflectors in order to alter the distance between specimen and objective, such as piezoelectric or galvanometer devices, in order to change this relationship. Translation or stage translation may cause changes; other mechanisms could involve piezoelectric sensors or galvanometer devices for greater control over this change.

Another scanning method utilizing oscillating mirrors controlled by servomotors consists of using one or more (servo-controlled) oscillating mirrors placed either horizontally or vertically, usually with low latency and variable speed z-axis scanning capability, providing free-line scanning capability of optical beam.

Isomet offers acousto-optic deflectors designed for both single-axis scanning and dual-axis scanning that can be driven with various radiofrequency (RF) frequencies. Each deflector features different open aperture sizes and resolution levels as well as drivers customized specifically to each type of acousto-optic deflector.

Input aperture

Laser scanning microscopy requires the use of acousto-optic deflectors to eliminate undesirable artifacts from laser light, while increasing optical resolution of specimens. Such deflectors eliminate interference caused by field and aperture diaphragms of microscopes as well as dirt that may accumulate on them or the light source.

A circular aperture in the focal plane of an objective lens serves to direct an expanded laser beam into the optical path of deflectors and redirect its light at various angles to scan samples in a raster pattern. This method is especially advantageous for point scanning as it enables users to access wide ranging data sets at suitable axial (z-step) intervals from just one specimen.

Acousto-optic deflectors can be made out of various crystals, with TeO2, GaP, LiNbO3 and ZnO being among the more frequently used ones [9]. These materials possess relatively high photoelastic tensor elements in certain crystallographic directions as well as excellent optical properties including low absorption rates, small refractive indices, uniaxial birefringence and low optical activity – essential characteristics of any deflector device.

Care must be taken in selecting materials and designing deflectors in order to reach maximum diffraction efficiency. This is particularly essential when designing continuous laser beam deflection – one or two axle axis – and vector (random) scanning applications where both lateral and axial scan ranges must be co-ordinated with each other.

Once a material has been selected, the next step should be determining the propagation direction and polarization of the diffracting acoustic mode. This step is pivotal as it determines an ideal power level necessary to guarantee maximum bandwidth from crystallographic modes.

Attaining this goal involves rotating the acoustic modes of deflectors by an angle related to their crystallographic optical axis and frequency in order to decrease chromatic dispersion due to acousto-optic interaction, known as an “acoustically rotated configuration”, via either an angular deviation of only the acoustic wave, or both acoustic and optical waves simultaneously.

Output aperture

Laser scanning microscopy uses an acousto-optic deflector to filter the optical beam, and reduce its intensity so as not to damage samples. This enables fast, ultrahigh resolution imaging as well as small sample imaging of up to 10nm samples.

Most acousto-optic deflectors adjust the refractive index of crystal by applying an ultrasonic alternating ultrasonic acoustic wave, traveling into it and creating periodic redistributions of refractive index. This causes part of incident laser light to deviate into an output beam used in microscope. Usually the acoustic component is tuned approximately inversely to its related laser frequency; transmission wavelength decreases with increasing frequency while extraordinary wavelength increases simultaneously.

Tuning relationships are crucial in determining diffraction efficiency of devices, which is defined as the product of the acoustic transit time across an aperture and frequency range width; an acoustic transit time refers to how long an acoustic wave takes to transit through a medium, which may differ for each deflector acousto-optic deflector.

To maximize diffraction efficiency, the phase between the acoustic wave and optical component of an output beam must be precisely aligned; this can be accomplished using acousto-optic gratings or Bragg diffusers.

Diffraction efficiency can also be affected by the angle range of an acoustic beam. As the wavelengths associated with acoustics can often extend further than optical wavelengths, usable output beam angular range tends to be relatively limited; to increase it further additional optical elements must be included.

Most commonly, acousto-optic tunable filters are constructed from TeO2 or other anisotropic crystals with good optical properties, such as antimony trioxide. These materials offer superior diffraction efficiency and low absorption across spectral ranges from the visible into near infrared regions; additionally they boast high acousto-optic figures, uniaxial birefringence, and small optical activity levels.

In order to maximize diffraction efficiency, the configuration of an acousto-optic deflector must be carefully considered when choosing its materials, shape optimization of active medium, and selection of surface roughness settings. This step in designing acousto-optic deflectors.
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