scispace - formally typeset
Search or ask a question
Journal ArticleDOI

Marker-free phase nanoscopy

01 Feb 2013-Nature Photonics (Nature Research)-Vol. 7, Iss: 2, pp 113-117
TL;DR: In this paper, a quasi-2 pi-holographic detection scheme and complex deconvolution was used to obtain a lateral resolution of 90 nm. But the results were limited to the case of living dendritic spines (neurones).
Abstract: We introduce a microscopic method that determines quantitative optical properties beyond the optical diffraction limit and allows direct imaging of unstained living biological specimens. In established holographic microscopy, complex fields are measured using interferometric detection, allowing diffraction-limited phase measurements. Here, we show that non-invasive optical nanoscopy can achieve a lateral resolution of 90 nm by using a quasi-2 pi-holographic detection scheme and complex deconvolution. We record holograms from different illumination directions on the sample plane and observe subwavelength tomographic variations of the specimen. Nanoscale apertures serve to calibrate the tomographic reconstruction and to characterize the imaging system by means of the coherent transfer function. This gives rise to realistic inverse filtering and guarantees true complex field reconstruction. The observations are shown for nanoscopic porous cell frustule (diatoms), for the direct study of bacteria (Escherichia coil), and for a time-lapse approach to explore the dynamics of living dendritic spines (neurones).
Citations
More filters
Journal ArticleDOI
TL;DR: This Review presents the main principles of operation and representative basic and clinical science applications of quantitative phase imaging, and aims to provide a critical and objective overview of this dynamic research field.
Abstract: Quantitative phase imaging (QPI) has emerged as a valuable method for investigating cells and tissues. QPI operates on unlabelled specimens and, as such, is complementary to established fluorescence microscopy, exhibiting lower phototoxicity and no photobleaching. As the images represent quantitative maps of optical path length delays introduced by the specimen, QPI provides an objective measure of morphology and dynamics, free of variability due to contrast agents. Owing to the tremendous progress witnessed especially in the past 10–15 years, a number of technologies have become sufficiently reliable and translated to biomedical laboratories. Commercialization efforts are under way and, as a result, the QPI field is now transitioning from a technology-development-driven to an application-focused field. In this Review, we aim to provide a critical and objective overview of this dynamic research field by presenting the scientific context, main principles of operation and current biomedical applications. Over the past 10–15 years, quantitative phase imaging has moved from a research-driven to an application-focused field. This Review presents the main principles of operation and representative basic and clinical science applications.

847 citations

Journal ArticleDOI
28 Mar 2013-Sensors
TL;DR: The principles of QPI are presented and some of the recent applications ranging from cell homeostasis to infectious diseases and cancer are highlighted, to provide important insight on how the QPI techniques potentially improve the study of cell pathophysiology.
Abstract: A cellular-level study of the pathophysiology is crucial for understanding the mechanisms behind human diseases. Recent advances in quantitative phase imaging (QPI) techniques show promises for the cellular-level understanding of the pathophysiology of diseases. To provide important insight on how the QPI techniques potentially improve the study of cell pathophysiology, here we present the principles of QPI and highlight some of the recent applications of QPI ranging from cell homeostasis to infectious diseases and cancer.

439 citations

Journal ArticleDOI
20 Feb 2015
TL;DR: In this article, the Fourier ptychography was used to estimate the 3D complex transmittance function of the sample at multiple depths, without any weak or single-scattering approximations.
Abstract: Realizing high resolution across large volumes is challenging for 3D imaging techniques with high-speed acquisition. Here, we describe a new method for 3D intensity and phase recovery from 4D light field measurements, achieving enhanced resolution via Fourier ptychography. Starting from geometric optics light field refocusing, we incorporate phase retrieval and correct diffraction artifacts. Further, we incorporate dark-field images to achieve lateral resolution beyond the diffraction limit of the objective (5× larger NA) and axial resolution better than the depth of field, using a low-magnification objective with a large field of view. Our iterative reconstruction algorithm uses a multislice coherent model to estimate the 3D complex transmittance function of the sample at multiple depths, without any weak or single-scattering approximations. Data are captured by an LED array microscope with computational illumination, which enables rapid scanning of angles for fast acquisition. We demonstrate the method with thick biological samples in a modified commercial microscope, indicating the technique’s versatility for a wide range of applications.

403 citations


Cites methods from "Marker-free phase nanoscopy"

  • ...2D phase retrieval methods are easily combined with tomography [4–12], but when wave-optical effects become more prominent (as in microscopy), diffraction tomography [13–19] becomes necessary....

    [...]

Journal ArticleDOI
TL;DR: An overview of cell refractive index models and measurement techniques including microfluidic chip-based techniques for the last 50 years are provided, the applications and significance of cellRefractive index in cell biology, hematology, and pathology are presented, and future research trends in the field are discussed.
Abstract: Cell refractive index is a key biophysical parameter, which has been extensively studied. It is correlated with other cell biophysical properties including mechanical, electrical and optical properties, and not only represents the intracellular mass and concentration of a cell, but also provides important insight for various biological models. Measurement techniques developed earlier only measure the effective refractive index of a cell or a cell suspension, providing only limited information on cell refractive index and hence hindering its in-depth analysis and correlation. Recently, the emergence of microfluidic, photonic and imaging technologies has enabled the manipulation of a single cell and the 3D refractive index of a single cell down to sub-micron resolution, providing powerful tools to study cells based on refractive index. In this review, we provide an overview of cell refractive index models and measurement techniques including microfluidic chip-based techniques for the last 50 years, present the applications and significance of cell refractive index in cell biology, hematology, and pathology, and discuss future research trends in the field, including 3D imaging methods, integration with microfluidics and potential applications in new and breakthrough research areas.

367 citations

Journal ArticleDOI
20 Jun 2015
TL;DR: A method for imaging 3D phase objects in a tomographic configuration implemented by training an artificial neural network to reproduce the complex amplitude of the experimentally measured scattered light is described.
Abstract: Optical tomography has been widely investigated for biomedical imaging applications. In recent years optical tomography has been combined with digital holography and has been employed to produce high-quality images of phase objects such as cells. In this paper we describe a method for imaging 3D phase objects in a tomographic configuration implemented by training an artificial neural network to reproduce the complex amplitude of the experimentally measured scattered light. The network is designed such that the voxel values of the refractive index of the 3D object are the variables that are adapted during the training process. We demonstrate the method experimentally by forming images of the 3D refractive index distribution of Hela cells.

340 citations


Cites methods from "Marker-free phase nanoscopy"

  • ...This idea was explored for conventional tomography in [22], and it could also be used in conjunction with the learning approach we describe in this paper....

    [...]

References
More filters
Journal ArticleDOI
TL;DR: Lateral resolution that exceeds the classical diffraction limit by a factor of two is achieved by using spatially structured illumination in a wide‐field fluorescence microscope with strikingly increased clarity compared to both conventional and confocal microscopes.
Abstract: Lateral resolution that exceeds the classical diffraction limit by a factor of two is achieved by using spatially structured illumination in a wide-field fluorescence microscope. The sample is illuminated with a series of excitation light patterns, which cause normally inaccessible high-resolution information to be encoded into the observed image. The recorded images are linearly processed to extract the new information and produce a reconstruction with twice the normal resolution. Unlike confocal microscopy, the resolution improvement is achieved with no need to discard any of the emission light. The method produces images of strikingly increased clarity compared to both conventional and confocal microscopes.

3,274 citations


"Marker-free phase nanoscopy" refers methods in this paper

  • ...In the final step, the reconstructed scattering potential in equation (1) was recast to real space by using equation (2),...

    [...]

Journal ArticleDOI
25 May 2007-Science
TL;DR: Initial applications indicate that emergent far-field optical nanoscopy will have a strong impact in the life sciences and in other areas benefiting from nanoscale visualization.
Abstract: In 1873, Ernst Abbe discovered what was to become a well-known paradigm: the inability of a lens-based optical microscope to discern details that are closer together than half of the wavelength of light. However, for its most popular imaging mode, fluorescence microscopy, the diffraction barrier is crumbling. Here, I discuss the physical concepts that have pushed fluorescence microscopy to the nanoscale, once the prerogative of electron and scanning probe microscopes. Initial applications indicate that emergent far-field optical nanoscopy will have a strong impact in the life sciences and in other areas benefiting from nanoscale visualization.

2,730 citations

Journal ArticleDOI
TL;DR: Off-axis holograms recorded with a magnified image of microscopic objects are numerically reconstructed in amplitude and phase by calculation of scalar diffraction in the Fresnel approximation to show that the transverse resolution is equal to the diffraction limit of the imaging system.
Abstract: We present a digital method for holographic microscopy involving a CCD camera as a recording device. Off-axis holograms recorded with a magnified image of microscopic objects are numerically reconstructed in amplitude and phase by calculation of scalar diffraction in the Fresnel approximation. For phase-contrast imaging the reconstruction method involves the computation of a digital replica of the reference wave. A digital method for the correction of the phase aberrations is presented. We present a detailed description of the reconstruction procedure and show that the transverse resolution is equal to the diffraction limit of the imaging system.

1,174 citations

PatentDOI
TL;DR: In this article, the authors present a system and methods for quantitative three-dimensional mapping of refractive index in living or non-living cells, tissues, or organisms using a phase-shifting laser interferometric microscope with variable illumination angle.
Abstract: The present invention relates to systems and methods for quantitative three-dimensional mapping of refractive index in living or non-living cells, tissues, or organisms using a phase-shifting laser interferometric microscope with variable illumination angle. A preferred embodiment provides tomographic imaging of cells and multicellular organisms, and time-dependent changes in cell structure and the quantitative characterization of specimen-induced aberrations in high-resolution microscopy with multiple applications in tissue light scattering.

844 citations

Journal ArticleDOI
TL;DR: The reconstruction algorithm is derived for parallel beam transmission computed tomography through two-dimensional structures in which diffraction of the insonifying beam must be taken into account and is applicable to diffraction tomography within either the first Born or Rytov approximations.

741 citations