Zichao Bian

Aperture-scanning Fourier ptychography for 3D refocusing and super-resolution macroscopic imaging

We report an imaging scheme, termed aperture-scanning Fourier ptychography, for 3D refocusing and super-resolution macroscopic imaging. The reported scheme scans an aperture at the Fourier plane of an optical system and acquires the corresponding intensity images of the object. The acquired images are then synthesized in the frequency domain to recover a high-resolution complex sample wavefront; no phase information is needed in the recovery process. We demonstrate two applications of the reported scheme. In the first example, we use an aperture-scanning Fourier ptychography platform to recover the complex hologram of extended objects. The recovered hologram is then digitally propagated into different planes along the optical axis to examine the 3D structure of the object. We also demonstrate a reconstruction resolution better than the detector pixel limit (i.e., pixel super-resolution). In the second example, we develop a camera-scanning Fourier ptychography platform for super-resolution macroscopic imaging. By simply scanning the camera over different positions, we bypass the diffraction limit of the photographic lens and recover a super-resolution image of an object placed at the far field. This platforms maximum achievable resolution is ultimately determined by the cameras traveling range, not the aperture size of the lens. The FP scheme reported in this work may find applications in 3D object tracking, synthetic aperture imaging, remote sensing, and optical/electron/X-ray microscopy.

Sparsely sampled Fourier ptychography

Fourier ptychography (FP) is an imaging technique that applies angular diversity functions for high-resolution complex image recovery. The FP recovery routine switches between two working domains: the spectral and spatial domains. In this paper, we investigate the spectral-spatial data redundancy requirement of the FP recovery process. We report a sparsely sampled FP scheme by exploring the sampling interplay between these two domains. We demonstrate the use of the reported scheme for bypassing the high-dynamic-range combination step in the original FP recovery routine. As such, it is able to shorten the acquisition time of the FP platform by ~50%. As a special case of the sparsely sample FP, we also discuss a sub-sampled scheme and demonstrate its application in solving the pixel aliasing problem plagued in the original FP algorithm. We validate the reported schemes with both simulations and experiments. This paper provides insights for the development of the FP approach.

Microscopy illumination engineering using a low-cost liquid crystal display

Illumination engineering is critical for obtaining high-resolution, high-quality images in microscope settings. In a typical microscope, the condenser lens provides sample illumination that is uniform and free from glare. The associated condenser diaphragm can be manually adjusted to obtain the optimal illumination numerical aperture. In this paper, we report a programmable condenser lens for active illumination control. In our prototype setup, we used a

InstantScope: a low-cost whole slide imaging system with instant focal plane detection.

15 liquid crystal display as a transparent spatial light modulator and placed it at the back focal plane of the condenser lens. By setting different binary patterns on the display, we can actively control the illumination and the spatial coherence of the microscope platform. We demonstrated the use of such a simple scheme for multimodal imaging, including bright-field microscopy, darkfield microscopy, phase-contrast microscopy, polarization microscopy, 3D tomographic imaging, and super-resolution Fourier ptychographic imaging. The reported illumination engineering scheme is cost-effective and compatible with most existing platforms. It enables a turnkey solution with high flexibility for researchers in various communities. From the engineering point-of-view, the reported illumination scheme may also provide new insights for the development of multimodal microscopy and Fourier ptychographic imaging.

Moisture-Responsive Wrinkling Surfaces with Tunable Dynamics

We report the development of a high-throughput whole slide imaging (WSI) system by adapting a cost-effective optomechanical add-on kit to existing microscopes. Inspired by the phase detection concept in professional photography, we attached two pinhole-modulated cameras at the eyepiece ports for instant focal plane detection. By adjusting the positions of the pinholes, we can effectively change the view angle for the sample, and as such, we can use the translation shift of the two pinhole-modulated images to identify the optimal focal position. By using a small pinhole size, the focal-plane-detection range is on the order of millimeter, orders of magnitude longer than the objectives depth of field. We also show that, by analyzing the phase correlation of the pinhole-modulated images, we can determine whether the sample contains one thin section, folded sections, or multiple layers separated by certain distances - an important piece of information prior to a detailed z scan. In order to achieve system automation, we deployed a low-cost programmable robotic arm to perform sample loading and

Single-frame rapid autofocusing for brightfield and fluorescence whole slide imaging

14 stepper motors to drive the microscope stage to perform x-y scanning. Using a 20X objective lens, we can acquire a 2 gigapixel image with 14 mm by 8 mm field of view in 90 seconds. The reported platform may find applications in biomedical research, telemedicine, and digital pathology. It may also provide new insights for the development of high-content screening instruments.

Rapid focus map surveying for whole slide imaging with continuous sample motion

The wrinkle dynamics (such as reversibility and stability) of human skin are affected by the external stimuli, as well as the skins structure and mechanical properties. Inspired by these tunable responses, three types of moisture-responsive wrinkle dynamics are achieved, for the first time, through a single film-substrate system. These dynamics include: (1) completely reversible wrinkles formation; (2) irreversible wrinkles formation I: the initially formed wrinkles can be permanently erased and never reappear; and (3) irreversible wrinkles formation II: once the wrinkles form, they can no longer be erased. The key to success is to control the stiffness and thickness ratios of the film and the substrate, and tailor the crosslink degree/gradient of the film to allow for moisture-dependent changes of modulus and swelling degree. These unique responsive dynamics motivate the invention of a series of optical devices triggered by moisture, including anticounterfeit tabs, encryption devices, water indicators, light diffusors, and antiglare films. This study also paves the road for further understanding of the skin wrinkling dynamics and manipulation.

Fourier ptychographic microscopy using wavelength multiplexing

A critical consideration for whole slide imaging (WSI) platform is to perform accurate autofocusing at high speed. Typical WSI systems acquire a z-stack of sample images and determine the best focal position by maximizing a figure of merit. This strategy, however, has suffered from several limitations, including low speed due to multiple image acquisitions, relatively low accuracy of focal plane estimation, short axial range for autofocusing, and difficulties in handling transparent samples. By exploring the autocorrelation property of the tissue sections, we report a novel single-frame autofocusing scheme to address the above challenges. In this approach, we place a two-pinhole-modulated camera at the epi-illumination arm. The captured image contains two copies of the sample separated by a certain distance. By identifying this distance, we can recover the defocus distance of the sample over a long z-range without z-scanning. To handle transparent samples, we set an offset distance to the autofocusing camera for generating out-of-focus contrast in the captured image. The single-frame nature of our scheme allows autofocusing even when the stage is in continuous motion. We demonstrate the use of the our autofocusing scheme for fluorescence WSI and quantify the focusing performance on 1550 different tissue tiles. The average autofocusing error is ~0.11 depth-of-field, 3 folds better than that of conventional methods. We report an autofocusing speed of 0.037 s per tile, which is much faster than that of conventional methods. The autofocusing range is ~80 µm, 8 folds longer than that of conventional methods. The reported scheme is able to solve the autofocusing challenges in WSI systems and may find applications in high-throughput brightfield/fluorescence WSI.

Stiffness analysis of 3D spheroids using microtweezers

Whole slide imaging (WSI) has recently been cleared for primary diagnosis in the U.S. A critical challenge of WSI is to perform accurate focusing in high speed. Traditional systems create a focus map prior to scanning. For each focus point on the map, a sample needs to be static in the x-y plane, and axial scanning is needed to maximize the contrast. Here we report a novel focus map surveying method for WSI. In this method, we illuminate the sample with two LEDs and recover the focus points based on 1D autocorrelation analysis. The reported method requires no axial scanning, no additional camera and lens, works for stained and transparent samples, and allows continuous sample motion in the surveying process. By using a 20× objective lens, we demonstrate a mean focusing error of ∼0.08  μm in the static mode and ∼0.17  μm in the continuous motion mode. The reported method may provide a turnkey solution for most existing WSI systems due to its simplicity, robustness, accuracy, and high speed. It may also standardize the imaging performance of WSI systems for digital pathology and find other applications in high-content microscopy, such as time-lapse live-cell imaging.

Dual light-emitting diode-based multichannel microscopy for whole-slide multiplane, multispectral and phase imaging

Abstract. Fourier ptychographic microscopy (FPM) is a recently developed technique stitching low-resolution images in Fourier domain to realize wide-field high-resolution imaging. However, the time-consuming process of image acquisition greatly narrows its applications in dynamic imaging. We report a wavelength multiplexing strategy to speed up the acquisition process of FPM several folds. A proof-of-concept system is built to verify its feasibility. Distinguished from many current multiplexing methods in Fourier domain, we explore the potential of high-speed FPM in spectral domain. Compatible with most existing FPM methods, our strategy provides an approach to high-speed gigapixel microscopy. Several experimental results are also presented to validate the strategy.