L09-Optical Microscopy1 PDF - Materials Characterization

MATL 5200/6200 Materials Characterization Optical or Light Microscopy Auburn University Materials Engineering  Some Types of Optical Microscopes 1. Simple optical microscope  One lens; 25x; 10 μm resolution 2. Stereoscopic microscope  Two lens trains; 20x to...

MATL 5200/6200 Materials Characterization Optical or Light Microscopy Auburn University Materials Engineering  Some Types of Optical Microscopes 1. Simple optical microscope  One lens; 25x; 10 μm resolution 2. Stereoscopic microscope  Two lens trains; 20x to 50x 3. Compound optical microscope  Objective + eyepiece + condenser lenses; 1300x; 1 μm resolution. Auburn University Materials Engineering  Optical Microscopy  1590 - Hans & Zacharias Janssen of Middleburg, Holland manufactured the first compound microscopes  Robert Hooke (1635-1703)- book Micrographia, published in 1665, devised the compound microscope most famous microscopical observation, which enabled variable focus and magnification to observe cells for the first time.  Most widely utilized method to examine microstructures of materials  Used in: – Quality control for materials processing, product development, etc. – Determining why a material failed – Establishing structure-property relationships in materials Auburn University Materials Engineering  Refraction  Provides the physical basis for optical lenses  Refraction: the change in direction of a wave due to a change in its transmission medium  Defined by refractive index (μ), a dimensionless number that describes how light, or any other radiation, propagates through that medium c Speed of light in a vacuum µ= = v Speed of light in the medium  where c is the speed of light in vacuum and v is the speed of light in the substance. https://micro.magnet.fsu.edu/primer/java/refra ction/refractionangles/index.html Auburn University Materials Engineering  Refraction He sees the fish here….. But it is really here!! Auburn University Materials Engineering  Refractive Index  For visible light most transparent media have refractive indices between 1 and 2  Diamond is very high, 2.42. This causes total internal reflection inside the diamond, The facets collect all the light and direct it out the top, so the diamond appears to glow or shine with extra light. Auburn University Materials Engineering  Effect of Refraction in Lens  Because the refracted angle αr depends on the incident angle αi, a convex lens can be used to focus light to a point at a specified distance from the lens.  The distance from the lens to that point is the principal focal length, f, of the lens.  Front and back faces don’t need to be symmetric.  Can further enhance by modifying μ outside the lens Auburn University Materials Engineering  Dispersion  Light with different wavelength (λ) refracts differently.  For example, violet light (λ ~ 400 nm) refracts more than red light (λ ~ 700 nm)  This is why we see rainbows www.rkm.com.au/.../animation-physics-prism.html http://www.xtal.iqfr.csic.es/Cristalo grafia/parte_05-en.html Auburn University Materials Engineering  Principles of Optical Microscopes  Image Formation by Convex Lens: The formation of an image when light rays, reflected from a mirror or refracted through a lens, converge to a point.  Magnification (M): Mismatching the focal length on either side of a lens or using 2 lenses of different focal length to change the size of the image in the virtual image. v− f M= f Auburn University Materials Engineering  Principles of Optical Microscopes Technically, its the ratio between image size to the object size. It can be varied by changing the distance between the object and the final lens (of the eye) or by inserting a second lens between the two. The magnification of a microscope can be calculated by linear optics, which tells us the magnification of a convergent lens M. Where f is the focal length of the lens and v is the distance between the image and lens. A higher magnification lens has a shorter focal length. Auburn University Materials Engineering  Principles of Optical Microscopes  When we examine microstructure with our eyes, the light path in a microscope goes through an eyepiece instead of projector lens to form a virtual image on the human eye.  Virtual image is usually taken as 25 cm from the eyepiece.  In modern microscopes, you can switch between eyepiece and projector lens for digital recording. Fig. 1.2 Schematic path of light in a microscope with eyepiece. Auburn University Materials Engineering  Resolution  When a point object is magnified, its image is a central spot (the Airy disk) surrounded by a series of diffraction rings Airy Patterns and the Limit of Resolution Auburn University Materials Engineering  Airy Disk https://www.youtube.com/watch?v=NazBRcMDOOo Auburn University Materials Engineering  Principles of Optical Microscopes  Resolution: The minimum distance between two points at which they can be visibly distinguished as two points. Resolution is controlled by the diffraction of light  In order to distinguish two point objects separated by a short distance, the Airy disks should not overlap each other. Controlling the size of the disc is what controls the resolution. The Airy disc size is also related to the wavelength of light. Thus the microscope resolution is: d 0.612λ R= = 2 µ sin α 0.612λ = NA https://micro.magnet.fsu.edu/primer/java/i mageformation/airydiskbasics/index.html Auburn University Materials Engineering  Numerical Aperture  The numerical aperture of a microscope objective is a measure of its ability to gather light and resolve fine specimen detail at a fixed object distance https://www.microscopyu.com/microscopy-basics/numerical-aperture Auburn University Materials Engineering  How to Improve Resolution  Reduce λ of light  Increase NA Auburn University Materials Engineering  Brightness & Contrast  Brightness is simply the intensity of light. It is related to both the Magnification and Numerical Aperture ( NA ) 2 Btransmission = M2  Contrast is defined as the relative change in light intensity (I) between an object and its background. In the case of microstructure we need contrast between the grains and the grain boundaries. I object − I background Contrast = I background Auburn University Materials Engineering  Depth of Field (DOF)  Depth of Field (DOF) is another important aspect. In optics the DOF is the portion of a scene that appears acceptably sharp in the image. Although a lens can precisely focus at only one distance, the decrease in sharpness is gradual on each side of the focused distance.  In optically-based microscopes the field of view is limited and when all of the object is not in the focal plane the out-of-plane areas will not be in focus.  Used in photography (f stop) Auburn University Materials Engineering  Depth of Field (DOF)  The same factors that effect resolution effect the depth of field but in the opposite way; therefore, a compromise must be reached between these two factors 1.22λ Df = µ sin α tan α Auburn University Materials Engineering  Depth of Field (DOF) Auburn University Materials Engineering  Depth of Focus  The range of image plane positions at which the image can be viewed without appearing out of focus for a fixed position of object. Often confused with depth of field but not the same. Not as important as depth of field. Depth of focus is M2 times depth of field. 1.22λ =Df ×M2 µ sin α tan α Auburn University Materials Engineering  Aberration  Aberration in optical systems (lenses intended to produce a sharp image) generally leads to blurring of the image. It occurs when light from one point of an object after transmission through the system does not converge into a single point.  Spherical (geometrical) aberration involves the light rays near the outer edges of the lens having different focal lengths. This is due to the curvature of the lens and is difficult to correct. This was the defect that the Hubble space telescope first had in 1990.  Chromatic aberration is caused by the dispersion of the lens material, the variation of its refractive index, n, with the wavelength of light, different wavelengths of light will be focused to different positions https://www.leica-microsystems.com/science-lab/eyepieces-objectives-and-optical-aberrations/ Auburn University Materials Engineering  Spherical aberration Chromatic aberration  Astigmatism : The off-axis image of a specimen point appears as a disc or blurred lines instead of a point.  Depending on the angle of the off-axis rays entering the lens, the line image may be oriented either tangentially or radially. Schematic of astigmatism caused by off-axis imaging and the appearance of the image at various distances https://www.microscopyu.com/ between the two focal planes, according to Hain et al. tutorials/astigmatism Auburn University Materials Engineering  Curvature of field (Coma aberration): The off-axis aberration occurs because the focal plane of an image is not flat but has a concave spherical surface http://olympus.magnet.fsu.edu/primer/techniques/confocal/confocalobjectives.html Auburn University Materials Engineering  How to reduce aberrations  Combine lenses with different shapes and μ’s to correct spherical and chromatic aberrations.  Select single λ illumination source to eliminate chromatic aberrations.  Less aberrations means more $$$. Auburn University Materials Engineering