October 5, 2006
Quantitative phase imaging and applications: a review
George R. Harrison
Spectroscopy Laboratory Massachusetts Institute of Technology,
Contributors: Takahiro Ikeda, YounKeun
Park, Niyom Lue, Wonshik Choi, Lauren Deflores, Kamran Badizadegan, Ramachandra
R. Dasari, and Michael S. Feld
Collaborators:
Abstract
We present a review of the quantitative phase imaging techniques
developed in the George R. Harrison Spectroscopy Laboratory at Massachusetts
Institute of Technology. The applications of these techniques to cell biology
are presented according to the time scales of the phenomena under investigation
. The work is shown in the broader context of quantitative phase imaging and
the research of other laboratories.
Table of contents
Review of quantitative phase imaging
3.
Full-field quantitative phase imaging
3.3. Diffraction phase microscopy
4.
Applications of full field techniques
4.1.1. Red
blood cell volumetry
4.2. Slow cell dynamics (minutes-days)
4.3 Rapid cell dynamics (milliseconds-seconds)
4.3.1.
Cell membrane fluctuations
4.3.2.
Fresnel particle tracking using diffraction phase microscopy
1.
Introduction
Phase contrast (PC) and differential interference contrast (DIC)
microscopy have been used extensively to infer morphometric features of live
cells without the need for exogenous contrast agents1. These
techniques transfer the information encoded in the phase of the imaging field
into the intensity distribution of the final image. Thus, the optical phase
shift through a given sample can be regarded as a powerful endogenous contrast
agent, as it contains information about both the thickness and refractive index
of the sample. However, both PC and DIC are qualitative
in terms of optical path-length measurement, i.e. the relationship between the
irradiance and phase of the image field is generally nonlinear2, 3.
Quantifying the optical phase shifts associated with cells gives access to
information about morphology and dynamics at the nanometer scale. Over the past
decade, the development of quantitative phase imaging techniques has received
increased scientific interest. The technology can be divided into single-point and full-field measurements, according to the experimental geometry
employed. Several point-measurement techniques have been applied for
investigating the structure and dynamics of live cells4-10. This type of measurement allows for fiber-optic implementation
and also high-speed punctual phase measurement by using a single, fast
photodetector. Full-filed phase measurement techniques, on the other hand,
provide simultaneous information from a large number of points on the sample,
which has the benefit of studying both the temporal and spatial behavior of the
biological system under investigation11-23.
In this paper, we review the main
quantitative phase imaging techniques and their applications reported in the
literature by the Spectroscopy Laboratory and others. The paper is organized as
follows.
2. Point measurements
The quantitative
retrieval of the phase delay associated with a biological sample offers
detailed information about the structure and its temporal evolution. This type
of information is not accessible through either amplitude-based microscopes (including optical coherence
tomography), or qualitative phase-based
techniques (such as phase contrast and interference contrast microscopy).
Various point-measurement
techniques have been developed over the years for quantifying phase shifts at a
given point through biological samples. This class of techniques can be
described as an extension of optical coherence tomography 24 to
measurements of phase, phase dispersion and birefringence associated with
biological structures. DeBoer et al. demonstrated depth-resolved birefringence
measurements with a polarization sensitive OCT system 25.
Differential phase-contrast OCT images have also generated with a
polarization-sensitive OCT instrument 26. Recently,
polarization-sensitive OCT was used to quantify phase retardation in the
retinal nerve fiber 27. An
instantaneous quadrature techniques was proposed based on using a 1xN fiber
coupler and the inherent phase shift between different output fibers 28.
Electrokinetic 29 and
thermo-refractive 30 properties
of tissue and tissue phantoms have been measured by differential phase OCT.
Phase sensitive OCT-type measurements have also been performed for studying
static cells 10, for
monitoring electric activity in nerves 8, 9, and spontaneous beating in cardiomyocytes 6.
The development of new technology for investigating biological structure and dynamics at the nanometer level has been a subject of intense research in the Low-coherence interferometry group at the Spectroscopy Laboratory. Thus, using a harmonically-related pair of wavelengths, a novel interferometer has been developed, which can provide high-stability phase dispersion information about transparent samples 5, 31. This technique was subsequently extended to the investigation of 3-dimensional objects; the new technique is referred to phase dispersion optical tomography