This PDF file contains the front matter associated with SPIE Proceedings Volume 8978, including the Title Page, Copyright information, Table of Contents, Introduction, and Conference Committee listing.

The speed of real-time adaptive optical systems is primarily restricted by the data processing hardware and computational aspects. Furthermore, the application of mirror layouts with increasing numbers of actuators reduces the bandwidth (speed) of the system and, thus, the number of applicable control algorithms. This burden turns out a key-impediment for deformable mirrors with continuous mirror surface and highly coupled actuator influence functions. In this regard, specialized hardware is necessary for high performance real-time control applications. Our approach to overcome this challenge is an adaptive optics system based on a Shack-Hartmann wavefront sensor (SHWFS) with a CameraLink interface. The data processing is based on a high performance Intel Core i7 Quadcore hard real-time Linux system. Employing a Xilinx Kintex-7 FPGA, an own developed PCie card is outlined in order to accelerate the analysis of a Shack-Hartmann Wavefront Sensor. A recently developed real-time capable spot detection algorithm evaluates the wavefront. The main features of the presented system are the reduction of latency and the acceleration of computation For example, matrix multiplications which in general are of complexity O(n³ are accelerated by using the DSP48 slices of the field-programmable gate array (FPGA) as well as a novel hardware implementation of the SHWFS algorithm. Further benefits are the Streaming SIMD Extensions (SSE) which intensively use the parallelization capability of the processor for further reducing the latency and increasing the bandwidth of the closed-loop. Due to this approach, up to 64 actuators of a deformable mirror can be handled and controlled without noticeable restriction from computational burdens.

Laser-induced mirror deformation and thermal lensing in optical high power systems shall be compensated by a thermally-piezoelectric deformable mirror (DM). In our device, the laser-induced thermal lensing is compensated by heating of the DM as previously described with compound loading. We experimentally show the capability of this mirror for wavefront shaping of up to 6.2 kW laser power and power densities of 2 kW/cm2. The laser-induced defocussing of the membrane is compensated by mirror heating. We introduce a new mirror setup with buried heater and temperature sensor elements. Therewith, the compensation of laser-induced mirror deformation is possible within the same time scale. The piezoelectric stroke of the single actuators depends on their position on the membrane, and is not affected by the reflected laser power.

For the passed several years, the Naval Research Laboratory (NRL) has been investigating the use of Carbon Fiber Reinforced Polymer (CFRP) material in the construction of a telescope assembly including the optical components. The NRL, Sandia National Laboratories (SNL), and Composite Mirror Applications, Inc. (CMA) have jointly assembled a prototype telescope and achieved “first light” images with a CFRP 0.4 m aperture telescope. CFRP offers several advantages over traditional materials such as creating structures that are lightweight and low coefficient of thermal expansion and conductivity. The telescope’s primary and secondary mirrors are not made from glass, but CFRP, as well. The entire telescope weighs approximately 10 kg while a typical telescope of this size would weigh quite a bit more. We present the achievement of “first light” with this telescope demonstrating the imaging capabilities of this prototype and the optical surface quality of the mirrors with images taken during a day’s quiescent periods.

Deformable mirrors, and particularly MEMS, are crucial components for the direct imaging of exoplanets for both ground-based and space-based instruments. Without deformable mirrors, coronagraphs are incapable of reaching contrasts required to image Jupiter-like planets. The system performance is limited by image quality degradation resulting from wavefront error introduced from multiple effects including: atmospheric turbulence, static aberrations in the system, non-common-path aberrations, all of which vary with time. Correcting for these effects requires a deformable mirror with fast response and numerous actuators having moderate stroke. Not only do MEMS devices fulfill this requirement but their compactness permits their application in numerous space- and ground-based instruments, which are often volume- and mass-limited. In this paper, I will briefly explain how coronagraphs work and their requirements. I then will discuss the Extreme Adaptive Optics needed to compensate for the introduced wavefront error and how MEMS devices are a good choice to achieve the performance needed to produce the contrasts necessary to detect exoplanets. As examples, I will discuss a facility instrument for the Gemini Observatory, called the Gemini Planet Imager, that will detect Jupiter-like planets and present recent results from the NASA Ames Coronagraph Experiment laboratory, in the context of a proposed space- based mission called EXCEDE. EXCEDE is planned to focus on protoplanetary disks.

We are developing an adaptive optics system for earth observing remote sensing sensor. In this system, high spatial resolution has to be achieved by a lightweight sensor system due to the launcher’s requirements. Moreover, simple hardware architecture have to be selected to achieve high reliability. Image based AOS realize these requirements without wavefront sensor. In remote sensing, it is difficult to use a reference point source unless the satellite controls its attitude toward a star. We propose the control algorithm of the deformable mirror on the basis of the extended scene instead of the point source.

Fluorescence fluctuation methods, such as fluorescence correlation spectroscopy, are very sensitive to optical aberrations. That is why it is possible to use a fluctuations-based metric, the molecular brightness, to correct aberrations using a sensorless modal adaptive optics approach. We have investigated the performance of this method by correcting known aberrations under various experimental conditions. The signal-to-noise ratio of the brightness measurement was examined theoretically and experimentally and found to be directly related to the accuracy of aberration correction, so that the latter can be predicted for a given sample brightness and measurement duration. We have also shown that the initial measurement conditions play a key role in the correction dynamics and we provide guidelines to optimize the corrections accuracy and speed. The molecular brightness, used as a metric, has the advantage that it depends on aberrations as the square of the Strehl ratio, regardless of the nature of the sample. Therefore, it is straightforward to predict the achievable correction accuracy and the same performance can be obtained in samples with different structure and contrast, which would not be possible with image-based optimization metrics.

Acquisition of images deep inside large samples is one of the most demanded improvements that current biology applications ask for. Absorption, scattering and optical aberrations are the main difficulties encountered in these types of samples. Adaptive optics has been imported form astronomy to deal with the optical aberrations induced by the sample. Nonlinear microscopy and SPIM have been proposed as interesting options to image deep into a sample. Particularly, light-sheet microscopy, due to its low photo bleaching properties, opens new opportunities to obtain information for example in long time lapses for large 3D imaging. In this work, we perform an overview of the application of adaptive optics to the fluorescence microscopy in linear and non-linear modalities. Then we will focus in the light-sheet microscopy architecture of two orthogonal optical paths which implies new requirements in terms of optical correction. We will see the different issues that appear in light-sheet microscopy particularly when imaging large and non-flat samples. Finally, we will study the problem of the isoplanetic patches.

The landscape of biomedical research in neuroscience has changed dramatically in recent years as a result of spectacular progress in dynamic microscopy. However, the optical accessibility of deep brain structures or deeper regions of the surgically exposed hippocampus (a few 100 microns typically) remains limited, due to volumic aberrations created by the sample inhomogeneities. Adaptive optics can correct for these aberrations. Our goal is to realize a novel adaptive optics module dedicated to in vivo two-photon calcium imaging of the hippocampus. The key issue in adaptive optics is the ability to perform an accurate and reliable wavefront sensing. In two- photon microscopy indirect methods are required. Two families of approaches have been proposed so far, the modal sensorless technique and a method based on pupil segmentation. We present here a formal comparison of these approaches, in particular as a function of the amount of aberrations.

A fast direct wavefront sensing method for dynamic in-vivo adaptive optical two photon microscopy has demonstrated. By using the direct wavefront sensing and open loop control, the system provides high-speed wavefront measurement and correction. To measure the wavefront in the middle of a Drosophila embryo at early stages, autofluorescence from endogenous fluorophores in the yolk were used as reference guide-stars. This method does not rely on fluorescently labeled proteins as guide-stars, which can simplify the sample preparation for wavefront measurement. The method was tested through live imaging of a Drosophila embryo. The aberration in the middle of the embryo was measured directly for the first time. After correction, both contrast and signal intensity of the structure in the middle of the embryo was improved.

The short focal length of the mouse eye gives rise to an optically thick retina (50 D). If in addition, multiple wavelengths are to be used simultaneously to image an arbitrary combination of retinal layers, the ≈ 10 D of longitudinal chromatic aberration means a total of 60 D of vergence must be covered. This dictates that marginal rays will cover a wide range of angles with respect to the optical axis at the pupil of a mouse (or murine) adaptive optics ophthalmoscope, in order to section through the entire retina with any wavelength simultaneously. In this work, we discuss the compromises associated with the design of a mouse adaptive optics ophthalmoscope using off-the-shelf spherical reflective and refractive optics.

The design of an Adaptive Optics (AO) Structured Illumination (SI) microscope is presented. Two key technologies are combined to provide effective super-resolution at significant depths in tissue. AO is used to measure and compensate for optical aberrations in both the system and the tissue by measuring the optical path differences in the wavefront. Uncorrected, these aberrations significantly reduce imaging resolution, particularly as we view deeper into tissue. SI allows us to reconstruct an image with resolution beyond the Rayleigh limit of the optics by aliasing high spatial frequencies, outside the limit of the optics, to lower frequencies within the system pass band. The aliasing is accomplished by spatially modulating the illumination at a frequency near the cutoff frequency of the system. These aliased frequencies are superimposed on the lower spatial frequencies of the object in our image. Using multiple images and an inverse algorithm, we separate the aliased and normal frequencies, restore them to their original frequency positions, and recreate the original spectrum of the object. This allows us to recreate a super-resolution image of the object. A problem arises with thick aberrating tissue. Tissue aberrations, including sphere, increase with depth into the tissue and reduce the high spatial frequency response of a system. This degrades the ability of SI to reconstruct at superresolution and limits its use to relatively shallow depths. However, adding AO to the system compensates for these aberrations allowing SI to work at maximum efficiency even deep within aberrating tissue.

μLinear Structured Illumination Microscopy (SIM) provides a two-fold increase over the diffraction limited resolution. SIM produces excellent images with 120nm resolution in tissue culture cells in two and three dimensions. For SIM to work correctly, the point spread function (PSF) and optical transfer function (OTF) must be known, and, ideally, should be unaberrated. When imaging through thick samples, aberrations will be introduced into the optical system which will reduce the peak intensity and increase the width of the PSF. This will lead to reduced resolution and artifacts in SIM images. Adaptive optics can be used to correct the optical wavefront restoring the PSF to its unaberrated state, and AO has been used in several types of fluorescence microscopy. We demonstrate that AO can be used with SIM to achieve 120nm resolution through 25m of tissue by imaging through the full thickness of an adult C. elegans roundworm. The aberrations can be corrected over a 25μm × 45μm field of view with one wavefront correction setting, demonstrating that AO can be used effectively with widefield superresolution techniques.

Stochastic Optical Reconstruction Microscopy (STORM) requires a high Strehl ratio point spread function (PSF) to achieve high resolution, especially in the presence of background fluorescence. The PSF is degraded by aberrations caused by imperfections in the optics, the refractive index mismatch between the sample and coverslip, and the refractive index variations of the sample. These aberrations distort the shape of the PSF and increase the PSF width directly reducing the resolution of STORM. Here we discuss the use of Adaptive Optics (AO) to correct aberrations, maintaining a high Strehl ratio even in thick tissue. Because the intensity fluctuates strongly from frame to frame, image intensity is not a reliable measure of PSF quality, and the choice of a robust optimization metric is critical. We demonstrate the use of genetic algorithms with single molecule imaging for optimization of the wavefront and introduce a metric that is relatively insensitive to image intensity. We demonstrate the correction of the wavefront from measurements of single quantum dots.

I will discuss our recent work on the use of digital optical phase conjugation and ultrasound tagging to accomplish timereversal deep tissue optical focusing for fluorescence imaging and other applications.