Structured illumination microscopy

Saturated Pattern Excitation or Saturated Structured Illumination Microscopy... 

M.G.L. Gustafsson, Nonlinear structured-illumination microscopy Wide-held huorescence imaging with theoretically unlimited resolution. Proc. Nat. Acad. Sci. U.S.A 102(37), 13081-13086 (2005)... 

Several far-field light microscopy methods have recently been developed to break the diffraction limit. These methods can be largely divided into two categories (1) techniques that employ spatially patterned illumination to sharpen the point-spread function of the microscope, such as stimulated emission depletion (STED) microscopy and related methods using other reversibly saturable optically linear fluorescent transitions (RESOLFT) [1,2], and saturated structured-illumination microscopy (SSIM) [3], and (2) a technique that is based on the localization of individual fluorescent molecules, termed Stochastic Optical Reconstruction Microscopy (STORM [4], Photo-Activated Localization Microscopy (PALM) [5], or Fluorescence Photo-Activation Localization Microscopy (FPALM) [6]. In this paper, we describe the concept of STORM microscopy and recent advances in the imaging capabilities of STORM. 

L. Schermelleh, P.M. Carlton, S. Haase, L. Shao, L. Winoto, P. Kner, B. Burke, M.C. Cardoso, D.A. Agard, M.G.L. Gustafsson, et ah. Subdiffraction multicolor imaging of the nuclear periphery with 3D structured illumination microscopy. Science 320, 1332-1336 (2008)... 

Fig. 6. Staining the actin cytoskeleton. Add 2x fixation buffer to plated cells and tlx for 20 min at 4°C. Remove fixation buffer and replace with stain buffer for 20 min protect from light. Shown Is an example of an FIL-60 cell stained with rhodamlne phalloldin visualized with structured Illumination microscopy.

Figure 6.15 Schematic representation of merozoites bound with nanomimics after egress from red blood cells (left). Projections of merozoites (blue) blocked by sulforhodamine B (SRB)-filled nanomimics observed with structured illumination microscopy (3D-SIM) and electron microscopy (EM) (right).

Figure 4. Fluorescence microscopy with axial resolution (A) TIRFM with an angle of illumination close to the critical angle c (B) laser scanning microscopy (C) structured illumination (D) image section calculated from structured illumination microscopy. U373-MG glioblastoma cells incubated with the mitochondrial maker rhodamine 123.

Figure 4 shows a comparison of fluorescence images of the mitochondrial marker rhodamine 123 when using the 3 microscopic techniques described above. Only those mitochondria which are close to the plasma membrane are detected by TIRFM (A), whereas mitochondria located in the middle of the cells are shown in the image sections selected by laser scanning microscopy (B) or calculated from structured illumination microscopy (D). 

Using saturated structured illumination, the nonlinearity stems from saturation of the excited state of fluorescent molecules once the illumination intensity is sufficient to excite most of the molecules, further increases in illumination intensity cannot yield corresponding increases in fluorescent emission. With this scheme, we have shown a point resolution of 50 nm, a further factor of two beyond what is achievable with normal, linear structured illumination microscopy. 

The cell distribution of 3-amino-4-hydroxymethyl acridine derivatives 70 (Scheme 25), which has the N3—C4—16 substitution pattern, was studied by real-time fluorescence microscopy and SIMS structured illumination microscopy). The latter method required the introduction of an iodine atom at position 6 of the acridine which influences the Upophihcity but does not modify significantly the biological properties such as IC50 and subcellular localization (2009EJMC4758). A co-polymer 71 consisting of water-soluble maleic anhydride-containing poly[maleic anhydride-u/i-acrylic acid] (poly(MA-alt-AA) or MAAA) copolymer was modified with acriflavine (AF) which displayed antibacterial activity on EHEC and Staphylococcus aureus (2014MI2903). 

Xu D, Jiang T, Li A et al (2013) Fast optical sectioning obtained by structured illumination microscopy using a digital mirror device. J Biomed Opt 18 060503... 

The gels contain inhomogeneities with a characteristic size of about 1 um Under favorable circumstances, this structure can be visualized in the wet gel by epi-illumination microscopy. When it can be visualized, the structure has a "spongy appearance, consistent with what one might expect of a system that has undergone spinodal decomposition. So far, however, there is no concrete evidence to support this mechanism of phase separation. 

Optical sectioning in wide-field microscopy may be achieved using the method of structured illumination. For example, a diffraction grating can be imaged in the plane of the sample, and the intensity resulting from this plane can be described by... 

The long-term goal of this project is to establish the feasibility and applicability of phytofluors as in vivo fluorescent probes. We also plan to investigate the development of switchable proteins for applications in the structured illumination approach to ultra-high resolution microscopy described above. 

PTM Photon tunneling microscopy [12] An interface is probed with an evanescent wave produced by internal reflection of the illuminating light Surface structure... 

Epi-illumination Subcellular imaging structures Freeze fracture Preparation of cellular ultrastructures in frozen-hydrated and living state for electron microscopy macromolecular organization of bilayer membranes... 

The structure (e.g., number, size, distribution) of fat crystals is difficult to analyze by common microscopy techniques (i.e., electron, polarized light), due to their dense and interconnected microstructure. Images of the internal structures of lipid-based foods can only be obtained by special manipulation of the sample. However, formation of thin sections (polarized light microscopy) or fractured planes (electron microscopy) still typically does not provide adequate resolution of the crystalline phase. Confocal laserscanning microscopy (CLSM), which is based on the detection of fluorescence produced by a dye system when a sample is illuminated with a krypton/argon mixed-gas laser, overcomes these problems. Bulk specimens can be used with CLSM to obtain high-resolution images of lipid crystalline structure in intricate detail. 

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