3-DIMENSIONAL Quantitative Phase Imaging and Microscopy of Bio and Non-Bio Transparent Phase Samples Using the Radial Shearing Interferometric Set-Up (original) (raw)

Quadriwave lateral shearing interferometry for quantitative phase microscopy of living cells

Optics Express, 2009

Phase imaging with a high-resolution wavefront sensor is considered. This is based on a quadriwave lateral shearing interferometer mounted on a non-modified transmission white-light microscope. The measurement technology is explained both in the scope of wave optics and geometrical optics in order to discuss its implementation on a conventional microscope. In particular we consider the effect of a non spatially coherent source on the phase-image signal-to-noise ratio. Precise measurements of the phase-shift introduced by microscopic beads or giant unilamellar vesicles validate the principle and show the accuracy of the methods. Diffraction limited images of living COS-7 cells are then presented, with a particular focus on the membrane and organelle dynamics.

Dynamic phase imaging of microscopic measurements using parallel interferograms generated from a cyclic shear interferometer

Optics Express, 2014

We present a technique which allows us to generate two parallel interferograms with phase shifts of π/2 using a Cyclic Shear Interferometer (CSI) and a polarizing splitter. Because of the use of a CSI, we obtain the derivative phase data map directly, due to its configuration, it is immune to vibrations because the reference wavefront and the object wavefront have a common path; the shearing interferometer is insensitive to temperature and vibration. To obtain the optical phase data map, two interferograms are generated by collocating a polarizing device at the output of the CSI. The optical phase was processed using a Vargas-Quiroga algorithm. Related experimental results obtained for dynamic microscopic transparent samples are presented.

Reflective interferometric chamber for quantitative phase imaging of biological sample dynamics

Journal of Biomedical Optics, 2010

We introduce a new interferometric setup for single-exposure wide-field holographic phase imaging of highly dynamic biological samples. In this setup, the interferometric signal originates from a specially designed reflective interferometric chamber ͑InCh͒, creating an off-axis interferogram on the output plane of the system. The setup only requires the InCh and a simple reflectionmode two lens imaging system, without the need for additional optical elements such as gratings in the beam path. In addition, due to the close-to-common-path geometry of the setup, phase noise is greatly reduced. We experimentally compare the inherent phase stability of the system in ambient conditions to that of a conventional interferometer. We also demonstrate use of this system for wide-field quantitative phase imaging of two different highly dynamic, optically transparent biological samples: beating myocardial cells and moving unicellular microorganisms.

Phase-shifting Zernike phase contrast microscopy for quantitative phase measurement

Optics Letters, 2011

Zernike phase contrast microscopy is extended and combined with a phase-shifting mechanism to perform quantitative phase measurements of microscopic objects. Dozens of discrete point light sources on a ring are constructed for illumination. For each point light source, three different levels of point-like phase steps are designed, which are alternatively located along a ring on a silica plate to perform phase retardation on the undiffracted (dc) component of the object waves. These three levels of the phase steps are respectively selected by rotating the silica plate. Thus, quantitative evaluation of phase specimens can be performed via phase-shifting mechanism. The proposed method has low "halo" and "shade-off" effects, low coherent noise level, and high lateral resolution due to the improved illumination scheme.

Dynamic phase imaging utilizing a 4-dimensional microscope system

Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XVIII, 2011

This paper describes a new, novel interference Linnik microscope system and presents images and data of live biological samples. The specially designed optical system enables instantaneous 4dimensional video measurements of dynamic motions within and among live cells without the need for contrast agents. This "label-free", vibration insensitive imaging system enables measurement of biological objects in reflection using harmless light levels with a variety of magnifications and wavelengths with fields of view from several hundred microns up to a millimeter. At the core of the instrument is a phase measurement camera (PMC) enabling simultaneous measurement of multiple interference patterns utilizing a pixelated phase mask taking advantage of the polarization properties of light. Utilizing this technology enables the creation of phase image movies in real time at video rates so that dynamic motions and volumetric changes can be tracked. Objects are placed on a reflective surface in liquid under a coverslip. Phase values are converted to optical thickness data enabling volumetric, motion and morphological studies. Data from a number of different organisms such as flagellates and rotifers will be presented, as will measurements of human breast cancer cells with the addition of various agents that break down the cells. These data highlight examples of monitoring different biological processes and motions.

Subsurface imaging and cell refractometry using quantitative phase/ shear-force feedback microscopy

20th International Conference on Optical Fibre Sensors, 2009

Over the last few years, several novel quantitative phase imaging techniques have been developed for the study of biological cells. However, many of these techniques are encumbered by inherent limitations including 2π phase ambiguities and diffraction limited spatial resolution. In addition, subsurface information in the phase data is not exploited. We hereby present a novel quantitative phase imaging system without 2 π ambiguities, which also allows for subsurface imaging and cell refractometry studies. This is accomplished by utilizing simultaneously obtained shear-force topography information. We will demonstrate how the quantitative phase and topography data can be used for subsurface and cell refractometry analysis and will present results for a fabricated structure and a malaria infected red blood cell.

Three-dimensional quantitative phase imaging: Current and future perspectives

Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing Xv, 2008

Great effort has been made in the recent past to develop new non-destructive imaging modalities for both two and three dimensional objects, based on the phase properties of a specimen. Quantitative phase tomography (QPT) is a hybrid technique that has been proposed to provide three-dimensional (3D) refractive index (RI) profiling of irregular phase objects by combining transverse phase measurements with traditional tomographic reconstruction techniques. This profiling is accomplished through measurements of sets of projections which are ultimately related to the RI values of the object's transverse cross-section. This is particularly useful for 3D refractive index determination of specimens where staining is not appropriate or for materials that cannot be stained and is essential to many applications in photonics and biotechnology. This article reviews recent developments in quantitative phase tomography as they are presently available and suggests future applications based on current research on the 3D RI. The enabling elements for 3D QPT in the context of four key areas are discussed: the effect of the refractive index of the surrounding matching fluid, spatial resolution, phase accuracy and optimal defocus. Recent progress and future perspectives related to each of these areas is presented with regard to various test objects of known optical properties.