Moreover, there is no assurance of a regular supply of high-quality cc-Kapton tape due to inconsistent industrial production procedures. Currently, the most commonly used tape for ATUM is carbon coated (cc)-Kapton tape (polyimide film, DuPont, Wilmington, USA) 5, 24, 32, but it has deficiencies due to a relatively high sheet resistance, non-uniform carbon coating that causes mottled surface resistance and scratches. Here we focus on the ATUM method, which allows for efficient, automated collection of thousands of serial ultrathin sections of uniform quality that subsequently can be imaged with SEM 24. EM volume datasets are typically obtained using methods, such as focused ion beam-scanning electron microscopy (FIB-SEM) 8, 19, 20, serial block-face electron microscopy (SBEM) 4, 21, 22, 23, automated tape-collecting ultramicrotomy (ATUM) with SEM 24, 25, 26, transmission electron microscope camera array (TEMCA) 7, 27, and transmission-mode SEM 28 in addition to conventional EM using ultramicrotomes with transmission electron microscopy (TEM) 6, 29, 30, with each method possessing unique benefits and drawbacks 31. Such structures include the entire nervous system 2, retina 3, 4, cortex 5, 6, 7, 8, myelin sheaths 9, endoplasmic reticulum 10, 11, renal pelvis 12, cornea 13, mitochondria 14, Drosophila brain 15, 16, plant tissue 17 and viral proteins 18. The electron microscopy (EM)-based reconstruction of neuronal circuits from serial ultrathin sections has attracted considerable recent attention, despite the emergence of super resolution microscopy 1, because EM is a reliable method for the diverse-scale analysis of dense nanoscale details in biological structures. We conclude that CNT tape can enable high-resolution volume electron microscopy for brain ultrastructure analysis. In addition, CNT tape is compatible with post-embedding immunostaining for light and electron microscopy. When combined with an enhanced en bloc staining protocol, CNT tape-processed brain sections reveal detailed synaptic ultrastructure. CNT tape can withstand multiple rounds of imaging, offer low surface resistance across the entire tape length and generate no wrinkles during the collection of ultrathin sections. Here we show that a plasma-hydrophilized carbon nanotube (CNT)-coated polyethylene terephthalate (PET) tape effectively resolves these issues and produces SEM images of comparable quality to those from transmission electron microscopy. Current tapes are limited by section wrinkle formation, surface scratches and sample charging during imaging. Section 9.Automated tape-collecting ultramicrotomy in conjunction with scanning electron microscopy (SEM) is a powerful approach for volume electron microscopy and three-dimensional neuronal circuit analysis. Triple Integrals in Cylindrical and Spherical Coordinates.Surfaces Defined Parametrically and Surface Area.Double Riemann Sums and Double Integrals over Rectangles. ![]() Constrained Optimization: Lagrange Multipliers.Directional Derivatives and the Gradient.Linearization: Tangent Planes and Differentials.10 Derivatives of Multivariable Functions.Derivatives and Integrals of Vector-Valued Functions.Functions of Several Variables and Three Dimensional Space.Active Calculus - Multivariable: our goals.
0 Comments
Leave a Reply. |