__FORCETOC__ Quantitative phase contrast microscopy or quantitative phase imaging are the collective names for a group of microscopy methods that quantify the

phase shift In physics and mathematics, the phase of a periodic function F of some real variable t (such as time) is an angle-like quantity representing the fraction of the cycle covered up to t. It is denoted \phi(t) and expressed in such a scale that it v ...

that occurs when light waves pass through a more optically dense object. Translucent objects, like a living human cell, absorb and scatter small amounts of light. This makes translucent objects much easier to observe in ordinary light microscopes. Such objects do, however, induce a phase shift that can be observed using a phase contrast microscope. Conventional phase contrast microscopy and related methods, such as

differential interference contrast microscopy Differential interference contrast (DIC) microscopy, also known as Nomarski interference contrast (NIC) or Nomarski microscopy, is an optical microscopy technique used to enhance the contrast in unstained, transparent samples. DIC works on the p ...

, visualize phase shifts by transforming phase shift gradients into intensity variations. These intensity variations are mixed with other intensity variations, making it difficult to extract quantitative information. Quantitative phase contrast methods are distinguished from conventional phase contrast methods in that they create a second so-called ''phase shift image'' or ''phase image'', independent of the intensity ( bright field) image.

Phase unwrapping Instantaneous phase and frequency are important concepts in signal processing that occur in the context of the representation and analysis of time-varying functions. The instantaneous phase (also known as local phase or simply phase) of a ''comple ...

methods are generally applied to the phase shift image to give absolute phase shift values in each pixel, as exemplified by Figure 1. Phase shift image of cells in 3D

The principal methods for measuring and visualizing phase shifts include

ptychography Ptychography (/t(ʌ)ɪˈkogræfi/ t(a)i-KO-graf-ee) is a computational method of microscopic imaging. It generates images by processing many coherent interference patterns that have been scattered from an object of interest. Its defining cha ...

and various types of holographic microscopy methods such as

digital holographic microscopy Digital holographic microscopy (DHM) is digital holography applied to microscopy. Digital holographic microscopy distinguishes itself from other microscopy methods by not recording the projected image of the object. Instead, the light wave fron ...

holographic interference microscopy Holographic interference microscopy (HIM) is holographic interferometry applied for microscopy for visualization of phase (waves), phase micro-objects. Phase micro-objects are invisible because they do not change Luminous intensity, intensity o ...

and digital in-line holographic microscopy. Common to these methods is that an

interference pattern In physics, interference is a phenomenon in which two waves combine by adding their displacement together at every single point in space and time, to form a resultant wave of greater, lower, or the same amplitude. Constructive and destructive ...

(

hologram Holography is a technique that enables a wavefront to be recorded and later re-constructed. Holography is best known as a method of generating real three-dimensional images, but it also has a wide range of other Holography#Applications, applic ...

) is recorded by a digital

image sensor An image sensor or imager is a sensor that detects and conveys information used to make an image. It does so by converting the variable attenuation of light waves (as they pass through or reflect off objects) into signals, small bursts of curr ...

. From the recorded interference pattern, the intensity and the phase shift image is numerically created by a computer

algorithm In mathematics and computer science, an algorithm () is a finite sequence of rigorous instructions, typically used to solve a class of specific Computational problem, problems or to perform a computation. Algorithms are used as specificat ...

. Quantitative phase contrast microscopy is primarily used to observed unstained living cells. Measuring the phase delay images of biological cells provides quantitative information about the morphology and the drymass of individual cells. Contrary to conventional phase contrast images, phase shift images of living cells are suitable to be processed by image analysis software. This has led to the development of non-invasive live cell imaging and automated

cell culture Cell culture or tissue culture is the process by which cells are grown under controlled conditions, generally outside of their natural environment. The term "tissue culture" was coined by American pathologist Montrose Thomas Burrows. This te ...

analysis systems based on quantitative phase contrast microscopy.published under CC BY 4.0 licensing
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References

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Phase shift time-lapse microscopy video of a triploid cell division

Cell identification with computational 3D holographic microscopy
{{Optical microscopy Microscopy Cell imaging Microbiology techniques Laboratory techniques Laboratory equipment Optical microscopy

See also

References

External links