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.
Scanning confocal microscopy is a standard choice for many fluorescence imaging applications in basic biomedical research. It is able to produce optically sectioned images and provide acquisition versatility to address many samples and application demands. However, scanning a focused point across the specimen limits the speed of image acquisition. As a result, scanning confocal microscope only works well with stationary samples. Researchers have performed parallel confocal scanning using digital-micromirror-device (DMD), which was used to project a scanning multi-point pattern across the sample. The DMD based parallel confocal systems increase the imaging speed while maintaining the optical sectioning ability. In this paper, we report the development of an add-on kit for high-speed and low-cost confocal microscopy. By adapting this add-on kit to an existing regular microscope, one can convert it into a confocal microscope without significant hardware modifications. Compared with current DMD-based implementations, the reported approach is able to recover multiple layers along the z axis simultaneously. It may find applications in wafer inspection and 3D metrology of semiconductor circuit. The dissemination of the proposed add-on kit under $1000 budget could also lead to new types of experimental designs for biological research labs, e.g., cytology analysis in cell culture experiments, genetic studies on multicellular organisms, pharmaceutical drug profiling, RNA interference studies, investigation of microbial communities in environmental systems, and etc.
The Fourier ptychography technique in reflection mode has great potential applications in tissue imaging and optical inspection, but the current configuration either has a limitation on cut-off frequency or is not practical. By placing the imaging aperture stop outside the illumination path, the illumination numerical aperture (NA) can be greater than the imaging NA of the objective lens. Thus, the cut-off frequency achieved in the proposed optical system is greater than twice the objective’s NA divided by the wavelength (2NAobj/λ), which is the diffraction limit for the cut-off frequency in an incoherent epi-illumination configuration. We experimentally demonstrated that the synthesized NA is increased by a factor of 4.5 using the proposed optical concept. The key advantage of the proposed system is that it can achieve high-resolution imaging over a large field of view with a simple objective. It will have a great potential for applications in endoscopy, biomedical imaging, surface metrology, and industrial inspection.
Circulating tumor cells (CTCs) are recognized as a candidate biomarker with strong prognostic and predictive potential in metastatic disease. Filtration-based enrichment technologies have been used for CTC characterization, and our group has previously developed a membrane microfilter device that demonstrates efficacy in model systems and clinical blood samples. However, uneven filtration surfaces make the use of standard microscopic techniques a difficult task, limiting the performance of automated imaging using commercially available technologies. Here, we report the use of Fourier ptychographic microscopy (FPM) to tackle this challenge. Employing this method, we were able to obtain high-resolution color images, including amplitude and phase, of the microfilter samples over large areas. FPM’s ability to perform digital refocusing on complex images is particularly useful in this setting as, in contrast to other imaging platforms, we can focus samples on multiple focal planes within the same frame despite surface unevenness. In model systems, FPM demonstrates high image quality, efficiency, and consistency in detection of tumor cells when comparing corresponding microfilter samples to standard microscopy with high correlation (R2=0.99932). Based on these results, we believe that FPM will have important implications for improved, high throughput, filtration-based CTC analysis, and, more generally, image analysis of uneven surfaces.
The on-chip detection of a weak optical signal in biological experiments can easily be complicated by the presence of an
overwhelming background signal, and as such, pre-detection background suppression is substantively important for
weak signal detection. In this paper, we report a structure that can be directly incorporated onto optical sensors to
accomplish background suppression prior to detection. This structure, termed surface-wave-enabled darkfield aperture
(SWEDA), consists of a central sub-wavelength hole surrounded by concentric grooves that are milled onto a gold layer.
Incoming light can be collected and converted into surface waves (SW) by the concentric grooves and then be recoupled
into propagating light through the central hole. We show that the SW-assisted optical component and the direct
transmission component of the central hole can cancel each other, resulting in near-zero transmission under uniform
illumination (observed suppression factor of 1230). This structure can therefore be used to suppress a light field's bright
background and allow sensitive detection of localized light field non-uniformity (observed image contrast enhancement
of 27dB). We also show that under a coherent background illumination, a CMOS pixel patterned with the proposed
structure achieves better SNR performance than an un-patterned single pixel.
We have developed a novel microscope technique that can achieve wide field-of-view (FOV) imaging and yet possess
resolution that is comparable to conventional microscope. The principle of wide FOV microscope system breaks the link
between resolution and FOV magnitude of traditional microscopes. Furthermore, by eliminating bulky optical elements
from its design and utilizing holographic optical elements, the wide FOV microscope system is more cost-effective. In
our system, a hologram was made to focus incoming collimated beam into a focus grid. The sample is put in the focal
plane and the transmissions of the focuses are detected by an imaging sensor. By scanning the incident angle of the
incoming beam, the focus grid will scan across the sample and the time-varying transmission can be detected. We can
then reconstruct the transmission image of the sample. The resolution of microscopic image is limited by the size of the
focus formed by the hologram. The scanning area of each focus spot is determined by the separation of the focus spots
and can be made small for fast imaging speed. We have fabricated a prototype system with a 2.4-mm FOV and 1-μm
resolution. The prototype system was used to image onion skin cells for a demonstration. The preliminary experiments
prove the feasibility of the wide FOV microscope technique, and the possibility of a wider FOV system with better resolution.