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==Techniques==
Single particle analysis can be done on both [[negative stain|negatively stained]] and vitreous ice-embedded [[Cryogenic transmission electron microscopy|CryoTEMcryomicroscopy]] (CryoTEM) samples. Single particle analysis methods are, in general, reliant on the sample being homogeneous, although techniques for dealing with [https://pubmed.ncbi.nlm.nih.gov/24089719/ conformational heterogeneity] are being developed.
Images (micrographs) are taken with an electron microscope using [[chargeCharge-coupled device|CCDcharged-coupled device]] (CCD) detectors coupled to a phosphorescent layer (in the past, they were instead collected on film and digitized using high-quality scanners). The image processing is carried out using specialized software [[Software tools for molecular microscopy|programs]], often run on multi-processor [[Computer cluster|computer clusters]]. Depending on the sample or the desired results, various steps of two- or three-dimensional processing can be done.
In addition, single particle analysis can also be performed in an individual particle mode using an ICP-MS unit.
===Image filtering===
Image filtering ([[band pass filter]]ing) is often used to reduce the influence of high and/or low [[spatial frequency]] information in the images, which can affect the results of the alignment and classification procedures. This is particularly useful in [[negative stain]] images. The algorithms make use of fast Fourier transforms ([[Fast Fourier transform|FFT]]), often employing [[Normal distribution|gaussianGaussian shaped]] soft-edged masks in [[Reciprocal lattice#Reciprocal space|reciprocal space]] to suppress certain frequency ranges. High-pass filters remove low spatial frequencies (such as ramp or gradient effects), leaving the higher frequencies intact. Low-pass filters remove high spatial frequency features and have a blurring effect on fine details.
===Contrast transfer function===
Due to the nature of image formation in the electron microscope, [[Bright-field microscopy|bright-field]] TEM images are obtained using significant [[Focus (optics)|underfocus]]. This, along with features inherent in the microscope's lens system, creates blurring of the collected images visible as a [[point spread function]]. The combined effects of the imaging conditions are known as the [[contrast transfer function]] (CTF), and can be approximated mathematically as a function in reciprocal space. Specialized image processing techniques such as phase flipping and amplitude correction / [[Wiener filter|wienerWiener filtering]] can (at least partially<ref name="Downing">{{Cite journal|vauthors=Downing KH, Glaeser RM |title=Restoration of weak phase-contrast images recorded with a high degree of defocus: the "twin image" problem associated with CTF correction |journal=Ultramicroscopy |volume=108 |issue=9 |pages=921–8 |date=August 2008 |pmid=18508199 |pmc=2694513 |doi=10.1016/j.ultramic.2008.03.004}}</ref>) correct for the CTF, and allow high resolution reconstructions.
===Three-dimensional reconstruction===
===Map visualization and fitting===
Various software [[Software tools for molecular microscopy|programs]] are available that allow viewing the 3D maps. These often enable the user to manually dock in protein coordinates (structures from [[X-ray crystallography]] or NMR) of subunits into the electron density. Several programs can also fit subunits computationally.
===Single particle ICP-MS===
SPSingle particle-ICP-MSinduced (Singlecoupled Particleplasma-Inducedmass Coupledspectroscopy Plasma(SP-Mass SpectroscopyICP-MS) is used in several areas where there is the possibility of detecting and quantifying suspended particles in samples of environmental fluids, assessing their migration, assessing the size of particles and their distribution, and also determining their stability in a given environment. Single Particle Inductively Coupled Plasma Mass Spectroscopy (SP -ICP-MS) was designed for particle suspensions in 2000 by Claude Degueldre. He first tested this new methodology at the Forel Institute of the University of Geneva and presented this new analytical approach at the 'Colloid 2oo2' symposium during the spring 2002 meeting of the EMRS, and in the proceedings in 2003[<ref>C. Degueldre & P. -Y. Favarger, « Colloid analysis by single particle inductively coupled plasma-mass spectroscopy: a feasibility study », Colloids and Surfaces A: Physicochemical and Engineering Aspects, symposium C of the E-MRS 2002 Spring Meeting in Strasbourg, France, vol. 217, no 1, 28 avril 2003, p. 137–142 (ISSN 0927-7757, DOI 10.1016/S0927-7757(02)00568-X)</ref>]. This study presents the theory of SP ICP-MS and the results of tests carried out on clay particles (montmorillonite) as well as other suspensions of colloids. This method was then tested on thorium dioxide nanoparticles by Degueldre & Favarger (2004),[<ref>C Degueldre et P. -Y Favarger, « Thorium colloid analysis by single particle inductively coupled plasma-mass spectrometry », Talanta, vol. 62, no 5, 19 avril 2004, p. 1051–1054 (ISSN 0039-9140, DOI 10.1016/j.talanta.2003.10.016</ref>] zirconium dioxide by Degueldre et al (2004)[<ref>C. Degueldre, P. -Y. Favarger et C. Bitea, « Zirconia colloid analysis by single particle inductively coupled plasma–mass spectrometry », Analytica Chimica Acta, vol. 518, no 1, 2 août 2004, p. 137–142 (ISSN 0003-2670, DOI 10.1016/j.aca.2004.04.015) </ref>] and gold nanoparticles, which are used as a substrate in nanopharmacy, and published by Degueldre et al (2006).[<ref>C. Degueldre, P. -Y. Favarger et S. Wold, « Gold colloid analysis by inductively coupled plasma-mass spectrometry in a single particle mode », Analytica Chimica Acta, vol. 555, no 2, 12 janvier 2006, p. 263–268 (ISSN 0003-2670, DOI 10.1016/j.aca.2005.09.021) </ref>] Subsequently, the study of uranium dioxide nano- and micro-particles gave rise to a detailed publication, Ref. Degueldre et al (2006).[<ref>C. Degueldre, P. -Y. Favarger, R. Rossé et S. Wold, « Uranium colloid analysis by single particle inductively coupled plasma-mass spectrometry », Talanta, vol. 68, no 3, 15 janvier 2006, p. 623–628 (ISSN 0039-9140, DOI 10.1016/j.talanta.2005.05.006, </ref>] Since 2010 the interest for SP ICP-MS has exploded.
==Examples==
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