Phase - contrast microscopy - wikipedia Phase - contrast microscopy Phase - contrast microscope A phase - contrast microscope Uses Microscopic observation of unstained biological material Inventor Frits Zernike Manufacturer Zeiss, Nikon, Olympus and others Related items Differential interference contrast microscopy, Hoffman modulation - contrast microscopy, Quantitative phase - contrast microscopy Phase - contrast microscopy is an optical microscopy technique that converts phase shifts in light passing through a transparent specimen to brightness changes in the image. Phase shifts themselves are invisible, but become visible when shown as brightness variations. When light waves travel through a medium other than vacuum, interaction with the medium causes the wave amplitude and phase to change in a manner dependent on properties of the medium. Changes in amplitude ( brightness ) arise from the scattering and absorption of light, which is often wavelength - dependent and may give rise to colors. Photographic equipment and the human eye are only sensitive to amplitude variations. Without special arrangements, phase changes are therefore invisible. Yet, phase changes often carry important information. The same cells imaged with traditional bright - field microscopy ( left ) and with phase - contrast microscopy ( right ) Phase - contrast microscopy is particularly important in biology. It reveals many cellular structures that are not visible with a simpler bright - field microscope, as exemplified in the figure. These structures were made visible to earlier microscopists by staining, but this required additional preparation and killed the cells. The phase - contrast microscope made it possible for biologists to study living cells and how they proliferate through cell division. After its invention in the early 1930s, phase - contrast microscopy proved to be such an advancement in microscopy that its inventor Frits Zernike was awarded the Nobel Prize in Physics in 1953. Working principle Play media Dark field and phase contrast microscopies operating principle The basic principle to making phase changes visible in phase - contrast microscopy is to separate the illuminating ( background ) light from the specimen - scattered light ( which makes up the foreground details ) and to manipulate these differently. The ring - shaped illuminating light ( green ) that passes the condenser annulus is focused on the specimen by the condenser. Some of the illuminating light is scattered by the specimen ( yellow ). The remaining light is unaffected by the specimen and forms the background light ( red ). When observing an unstained biological specimen, the scattered light is weak and typically phase - shifted by − 90 ° ( due to both the typical thickness of specimens and the refractive index difference between biological tissue and the surrounding medium ) relative to the background light. This leads to the foreground ( blue vector ) and background ( red vector ) having nearly the same intensity, resulting in low image contrast. In a phase - contrast microscope, image contrast is increased in two ways : by generating constructive interference between scattered and background light rays in regions of the field of view that contain the specimen, and by reducing the amount of background light that reaches the image plane. First, the background light is phase - shifted by − 90 ° by passing it through a phase - shift ring, which eliminates the phase difference between the background and the scattered light rays. When the light is then focused on the image plane ( where a camera or eyepiece is placed ), this phase shift causes background and scattered light rays originating from regions of the field of view that contain the sample ( i.e., the foreground ) to constructively interfere, resulting in an increase in the brightness of these areas compared to regions that do not contain the sample. Finally, the background is dimmed ~ 70 - 90 % by a gray filter ring -- this method maximizes the amount of scattered light generated by the illumination ( i.e., background ) light, while minimizing the amount of illumination light that reaches the image plane. Some of the scattered light ( which illuminates the entire surface of the filter ) will be phase - shifted and dimmed by the rings, but to a much lesser extent than the background light ( which only illuminates the phase - shift and gray filter rings ). The above describes negative phase contrast. In its positive form, the background light is instead phase - shifted by + 90 °. The background light will thus be 180 ° out of phase relative to the scattered light. The scattered light will then be subtracted from the background light to form an image with a darker foreground and a lighter background, as shown in the first figure. Related methods Further information : differential interference contrast microscopy, hoffman modulation contrast microscopy, and quantitative phase - contrast microscopy Cultured cells imaged by DIC microscopy A quantitative phase - contrast microscopy image of cells in culture. The height and color of an image point correspond to the optical thickness, which only depends on the object 's thickness and the relative refractive index. The volume of an object can thus be determined when the difference in refractive index between the object and the surrounding media is known. The success of the phase - contrast microscope has led to a number of subsequent phase - imaging methods. In 1952 Georges Nomarski patented what is today known as differential interference contrast ( DIC ) microscopy. It enhances contrast by creating artificial shadows, as if the object is illuminated from the side. But DIC microscopy and phase contrast microscopy both use polarized light, which is unsuitable when the object or its container alter polarization. With the growing use of polarizing plastic containers in cell biology, DIC microscopy is increasingly replaced by Hoffman modulation contrast microscopy, invented by Robert Hoffman in 1975. Traditional phase - contrast methods enhance contrast optically, blending brightness and phase information in single image. Since the introduction of the digital camera in the mid-1990s, several new digital phase - imaging methods have been developed, collectively known as quantitative phase - contrast microscopy. These methods digitally create two separate images, an ordinary bright - field image and a so - called phase - shift image. In each image point, the phase - shift image displays the quantified phase shift induced by the object, which is proportional to the optical thickness of the object. See also Live cell imaging Phase - contrast imaging Phase - contrast X-ray imaging References Jump up ^ `` The phase contrast microscope ''. Nobel Media AB. Jump up ^ Zernike, F. ( 1955 ). `` How I Discovered Phase Contrast ''. Science. 121 ( 3141 ) : 345 -- 349. Bibcode : 1955Sci... 121... 345Z. doi : 10.1126 / science. 121.3141. 345. PMID 13237991. Jump up ^ `` The Nobel Prize in Physics 1953 ''. Nobel Media AB. Jump up ^ `` Phase contrast microscopy ''. Phase Holographic Imaging AB. Jump up ^ `` Introduction to Phase Contrast Microscopy ''. Nikon MicroscopyU. Jump up ^ `` Phase Contrast ''. Leica Science Lab. Jump up ^ Frits Zernike ( 1942 ). `` Phase contrast, a new method for the microscopic observation of transparent objects part I ''. Physica. 9 ( 7 ) : 686 -- 698. Bibcode : 1942Phy... 9... 686Z. doi : 10.1016 / S0031 - 8914 ( 42 ) 80035 - X. Jump up ^ Frits Zernike ( 1942 ). `` Phase contrast, a new method for the microscopic observation of transparent objects part II ''. Physica. 9 ( 10 ) : 974 -- 980. Bibcode : 1942Phy... 9... 974Z. doi : 10.1016 / S0031 - 8914 ( 42 ) 80079 - 8. Jump up ^ Oscar Richards ( 1956 ). `` Phase Microscopy 1954 - 56 ''. Science. 124 ( 3226 ) : 810 -- 814. Bibcode : 1956Sci... 124... 810R. doi : 10.1126 / science. 124.3226. 810. Jump up ^ US2924142, Georges Nomarski, `` INTERFERENTIAL POLARIZING DEVICE FOR STUDY OF PHASE OBJECTS '' Jump up ^ US4200354, Robert Hoffman, `` Microscopy systems with rectangular illumination particularly adapted for viewing transparent object '' Jump up ^ Kemmler, M. ; Fratz, M. ; Giel, D. ; Saum, N. ; Brandenburg, A. ; Hoffmann, C. ( 2007 ). `` Noninvasive time - dependent cytometry monitoring by digital holography ''. Journal of Biomedical Optics. 12 ( 6 ) : 064002. Bibcode : 2007JBO... 12f4002K. doi : 10.1117 / 1.2804926. PMID 18163818. Jump up ^ `` Quantitative phase contrast microscopy ''. Phase Holographic Imaging AB. External links Optical Microscopy Primer -- Phase Contrast Microscopy by Florida State University Phase contrast and dark field microscopes ( Université Paris Sud ) Library resources about Phase - contrast microscopy Resources in your library Resources in other libraries Wikimedia Commons has media related to Phase contrast microscopic images. Optical microscopy Microscope Optical microscopy Illumination and contrast methods Bright - field microscopy Köhler illumination Dark - field microscopy Phase contrast Quantitative phase - contrast microscopy Differential interference contrast ( DIC ) Dispersion staining Second harmonic imaging ( SHIM ) 4Pi microscope Structured illumination Sarfus Fluorescence methods Fluorescence microscopy Confocal microscopy Two - photon excitation microscopy Multiphoton microscopy Image deconvolution Total internal reflection fluorescence microscopy ( TIRF ) Lightsheet microscopy ( LSFM / SPIM ) Sub-diffraction limit techniques Diffraction limit Stimulated emission depletion ( STED ) Photo - activated localization microscopy ( PALM / STORM ) Near - field ( NSOM / SNOM ) Retrieved from `` https://en.wikipedia.org/w/index.php?title=Phase-contrast_microscopy&oldid=837437694 '' Categories : Microscopy Dutch inventions Hidden categories : Commons category with local link different than on Wikidata Talk About Wikipedia Català Deutsch Español Français Кыргызча Magyar Nederlands 日本 語 Norsk Polski Português Русский Slovenščina Українська 中文 7 more Edit links This page was last edited on 20 April 2018, at 20 : 25. 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Question: where would phase contrast microscopy improve image quality
Answer: cellular structures