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McGill University
Anatomy & Cell Biology
ANAT 262
Isabelle Rouiller

ndANAT 262 January 22 2013 I Rouiller Lecture 05 ANAT 262 Lecture 4Electron MicroscopyndIsabelle Rouiller January 22 2013NOTE This NTC is meant to be used as a study aid to supplement your own class notes Hence not all of the text contained in the lecture slides will be reproduced herePlease send comments or questions about NTCs to us through email macssvpacademicgmailcom Announcements MACSS office is currently under construction Any questions or concerns should be sent to macssu1repsgmailcom ReferencesMolecular Biology of the Cell 5th Edition B Alberts et al pp604613 Uses of Electron Microscopy EMStudying tissue morphology and changes in morphology caused by various diseases ie change in ribosomal morphology may be evident of cellular mutation under EM such as abnormally large lysosome sizeObserving protein localization changes at high resolution ie trafficking among cellular compartments at specified times in normal cells Studying structural biology looking at molecules in order to determine their 3D structure to then further understand how they function began in the 1960s1 ndANAT 262 January 22 2013 I Rouiller Lecture 05 Resolving Power of EMResolving power is defined as the ability of the equipment to discern two objects in close proximity and the distance between them Thus resolving power is a property of an instrument in use The main advantage that electron microscopy has over light microscopy is the fact that the resolving power of the EM 05A is far greater than that of a light microscope 200 nm2000A Greater than or equal to the actual resolution usually far better Resolution is a dependent on the experiment while resolving power is a constant that is dependent on the equipment in use In other words resolution is a measure of the experiment while resolving power is a property of the instrument in use A resolution of 05A the value of the resolving power for EM has not yet been attained in electron microscopy This is because biological samples are difficult to work with The best resolution attained with EM for a biological sample is 2A and for this reason there is much room for improvement within the instrument Resolution The limit of resolution d is the smallest distance between two points that can still be distinguished This property is used widely in all microscopy Based on optics resolution may be calculated using the following equationhalf the angular width of the cone of rays collected by the objective lens from a typical point in the specimen the greater this value the lower the value of resolution but since the maximum width is 180 sinhas a maximum value of 1 to improve resolutionnthe refractive index of the medium usually air for light microscopy or oil for electron microscopy separating the specimen from the objective and condenser lensesthe wavelength of light usedObjective lens collects a cone of light rays to create a magnified imageCondenser lens focuses a cone of light rays onto each point in the specimenIn order to obtain better results using EM the resolution should be as small a value as possible The only possible way to reduce the resolution is to utilize smaller wavelengths The wavelengths used in light microscopy all exist within the visible light spectrum of 400800 nm For this reason the resolution of light microscopy is limited to the range of these values The use of electrons in EM allow for much small wavelengths 2
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