Nanoscopy with focused light
... Throughout the 20th century it was widely accepted that a light microscope relying on
conventional optical lenses cannot discern details that are much finer than about half
the wavelength of light (200-400 nm), due to diffraction. However, in the 1990s, the
viability to overcome the diffraction barr ...
High Resolution Biomedical Imaging with Light and Sound
... High resolution imaging systems are invaluable tools for biomedical research and clinical
practice. Photoacoustic microscopy (PAM) is an emerging hybrid technique that can overcome
the limitations of conventional optical and ultrasonic imaging modalities. A pulsed laser
illuminates tissue, where opt ...
... Most biological objects, however, absorb only weakly in the
visible spectrum. This lead to:
• Development of specific staining (nowadays almost entirely
replaced by fluorescent labeling)
• Development of UV microscopy (Köhler) facing technical
difficulties due to absorption of UV light by glass
• Us ...
... and useful explanations are included. The terms appear in
bold lettering and many interact with other terms in bold lettering. Note that many of these variables are usually thought
of in terms of their effect on spatial resolution. They are listed
here because reduced resolution translates into “put ...
Biology 177: Principles of Modern Microscopy
... We have looked at several different methods for optical sectioning
of fluorescent samples. The two main methods are Laser Scanning
Confocal Microscopy (LSCM) and light sheet microscopy or
Selective Plane Illumination Microscopy (SPIM). LSCM has been
around a long time compared to SPIM.
Question: Do ...
Compensated lens-free light field microscopy
... Lens-free light field microscopy (LLFM) is a holographic microscope with large field of view
which produces an image of micro-range resolution using an interferometric reconstruction
method . The quality of LLFM imaging is limited due to the effect of image degradation
factors such as (1) optical ...
Prof. Knut W. Urban
... - An expedition into the world of atoms by aberration-corrected electron optics
The realization of aberration-corrected lenses has triggered a quantum jump in electron optics. The recent
generation of transmission electron microscopes with aberration-corrected optics allows materials science in
Measurement of the 4Pi-confocal point spread function proves 75
... Cambridge, England). The bead was scanned 1.5 pm along
the optical axis and 0.8 pm in lateral direction. The smallest
resolvable step of the stage was 10 nm, thus providing a
high-precision measurement of the PSF.
The confocal resolution was determined by using the
4Pi-confocal arrangement with the ...
super-resolved fluorescence microscopy
... principle can also be implemented by Saturated Structural Illumination Microscopy (SSIM)
(Gustafsson, 2005). These methods can be used when the fluorescing regions contain ensembles
of fluorophores as well as single fluorophores and are therefore generically referred to as
“Super-resolved ensemble f ...
... Modulation transfer function
• Resolution and
performance of optical
microscope can be
characterized by the
• MTF is measurement of
microscope's ability to
transfer contrast from
the specimen to the
image plane at specific
• Incorporates resolution
Super-resolution microscopy is a form of light microscopy. Due to the diffraction of light, the resolution of conventional light microscopy is limited as stated by Ernst Abbe in 1873. A good approximation of the resolution attainable is the full width at half maximum (FWHM) of the point spread function, and a precise widefield microscope with high numerical aperture and visible light usually reaches a resolution of ~250 nm.Super-resolution techniques allow the capture of images with a higher resolution than the diffraction limit. They fall into two broad categories, ""true"" super-resolution techniques, which capture information contained in evanescent waves, and ""functional"" super-resolution techniques, which use clever experimental techniques and known limitations on the matter being imaged to reconstruct a super-resolution image.True subwavelength imaging techniques include those that utilize the Pendry Superlens and near field scanning optical microscopy, the 4Pi Microscope and structured illumination microscopy technologies like SIM and SMI. However, the majority of techniques of importance in biological imaging fall into the functional category.There are two major groups of methods for functional super-resolution microscopy: Deterministic super-resolution: The most commonly used emitters in biological microscopy, fluorophores, show a nonlinear response to excitation, and this nonlinear response can be exploited to enhance resolution. These methods include STED, GSD, RESOLFT and SSIM. Stochastic super-resolution: The chemical complexity of many molecular light sources gives them a complex temporal behaviour, which can be used to make several close-by fluorophores emit light at separate times and thereby become resolvable in time. These methods include SOFI and all single-molecule localization methods (SMLM) such as SPDM, SPDMphymod, PALM, FPALM, STORM and dSTORM.On October 8th, 2014, the Nobel Prize in Chemistry was awarded to Eric Betzig, W.E. Moerner and Stefan Hell for ""the development of super-resolved fluorescence microscopy,"" which brings ""optical microscopy into the nanodimension"".