Direct laser writing (DLW) has been widely used in a variety of engineering and research applications. However, the fabrication of complex and robust three-dimensional (3D) structures at submicron-level resolution by DLW is still largely limited by the laser focus quality, i.e., point spread function (PSF), laser dose, precision of mechanical scanners, and printing trajectory. In this work, we present a two-photon polymerization (TPP)-based DLW system based on a digital micromirror device (DMD) and binary holography to realize aberration-free large-area stitch-free 3D printing as well as 3D random-access scanning. First, the binary holograms, which control the amplitude, phase, and position of the laser focus, are optimized by the sensorless adaptive optics algorithm to correct the distorted wavefront in the DMD work field. Next, the DMD is synchronized to a continuously moving sample stage to eliminate stitching errors, i.e., the sample positioner simultaneously moves with the scanning focus until the structure is completed. We have fabricated large-area complex 3D structures, e.g., metamaterial structures, and micro-lenses, and 2D gray level diffractive optical elements (DOEs) with better than 100 nm resolution and optimal scanning trajectories. Notably, the variation of the scanning trajectory, laser power (dose), and voxel sizes can be realized without affecting the scanning speed, i.e., 22.7 kHz, which is equivalent to the DMD pattern rate.
In this Letter, we present a new, to our knowledge, aberration-free 3D imaging technique based on digital micromirror device (DMD)-based two-photon microscopy and sensorless adaptive optics (AO), where 3D random-access scanning and modal wavefront correction are realized using a single DMD chip at 22.7 kHz. Specifically, the DMD is simultaneously used as a deformable mirror to modulate a distorted wavefront and a fast scanner to maneuver the laser focus in a 3D space by designed binary holograms. As such, aberration-free 3D imaging is realized by superposing the wavefront correction and 3D scanning holograms. Compared with conventional AO devices and methods, the DMD system can apply optimal wavefront correction information to different imaging regions or even individual pixels without compromising the scanning speed and device resolution. In the experiments, we first focus the laser through a diffuser and apply sensorless AO to retrieve a corrected focus. After that, the DMD performs 3D scanning on a Drosophila brain labeled with green fluorescent protein. The two-photon imaging results, where optimal wavefront correction information is applied to 3 × 3 separate regions, demonstrate significantly improved resolution and image quality. The new DMD-based imaging solution presents a compact, low-cost, and effective solution for aberration-free two-photon deep tissue imaging, which may find important applications in the field of biophotonics.
We present the modular design and characterization of a multi-modality video-rate two-photon excitation (TPE) microscope based on integrating a digital micromirror device (DMD), which functions as an ultrafast beam shaper and random-access scanner, with a pair of galvanometric scanners. The TPE microscope system realizes a suite of new imaging functionalities, including (1) multi-layer imaging with 3D programmable imaging planes, (2) DMD-based wavefront correction, and (3) multi-focus optical stimulation (up to 22.7 kHz) with simultaneous TPE imaging, all in real-time. We also report the detailed optomechanical design and software development that achieves high level system automation. To verify the performance of different microscope functions, we have devised and performed imaging experiments on Drosophila brain, mouse kidney and human stem cells. The results not only show improved imaging resolution and depths via the DMD-based adaptive optics, but also demonstrate fast multi-focus stimulation for the first time. With the new imaging capabilities, e.g., tools for optogenetics, the multi-modality TPE microscope may play a critical role in the applications pertinent to neuroscience and biophotonics.
A system for making wavefront corrections for use in multiphoton microscopy has been constructed. Corrections are made using a high-resolution nematic liquid crystal device which has a phase stroke of 2π. The device has a design wavelength of 1064 nm. A simple way for setting the device up for lower wavelengths (here 800 nm) is presented. It was found that the device has an undesired zero-order diffraction component of 30%. A scheme for filtering this portion out is presented and it was demonstrated that this can eliminate the component completely. The device was used to optically simulate a thin lens with a specified focal length, which was found to match within error bounds. Finally the modulator was used to compensate for a mechanical defocus that was applied intentionally.
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