Chapter 7 Companion site for Light and Video Microscopy Author: Wayne

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 2 Common (A,B), partially-polarized (C) and elliptically- (D), circularly- (E) and linearly- (F,G) polarized light. FIGURE 7.1

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 3 The resultant of two coherent waves is determined by finding the vector sum of the components. Moreover, any single wave can be viewed as being composed of two orthogonal components. This is the double reality of vectors and light. FIGURE 7.2

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 4 Demonstration of transverse waves with a rope or long spring. FIGURE 7.3

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 5 Herapath discovered that the crystals that formed in the iodine-stained urine of a dog that had been fed quinine were either bright or dark in the places where they overlapped. Since their transparency or opacity of the thin crystals depended on their mutual orientation, he realized that they produced polarized light and he had discovered that the most powerful polarizing substance known … proved to be a new salt of a vegetable alkaloid. FIGURE 7.4

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 6 Bartholinus noticed that the light transmitted through a piece of calcite formed two images and must therefore be split into two beams. FIGURE 7.5

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 7 Huygens explained double refraction in terms of wave theory. He proposed that the atoms in the calcite acted asymmetrically upon the incoming light so that the waves that made up the ordinary beam were spherical and passed straight through the crystal, whereas the waves that made up the extraordinary beam were elliptical and traveled diagonally through the crystal. If he rotated the crystal while he viewed the double image through the top of crystal, the image produced by the ordinary ray (o) would remain stationary while the image produced by the extraordinary ray would precess around the ordinary image. FIGURE 7.6

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 8 Production of polarized light by reflection at Brewsters angle. According to Malus, the plane of polarization was the plane that included the incident ray and the reflected ray. FIGURE 7.7

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 9 Production of polarized light by scattering sunlight from the molecules in the atmosphere. FIGURE 7.8

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 10 Extinguishing polarized light using a polarizer whose azimuth of maximal transmission is perpendicular to the azimuth of the linearly polarized light formed by the polarizer. FIGURE 7.9

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 11 A polarimeter. FIGURE 7.10

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 12 The design of and conventions used in a polarized light microscope. FIGURE 7.11

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 13 (A) Common light being split by calcite and (B) by a Nicol prism formed from two pieces of calcite cemented together with Canada balsam. FIGURE 7.12

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 14 Studying the interaction of polarized light with electrons using microwave radiation. FIGURE 7.13

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 15 Propagation of waves through negatively and positively birefringent materials when the incident light strikes parallel, perpendicular, and oblique to the optic axis. Ordinary waves (), extraordinary waves ( ). FIGURE 7.14

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 16 Determination of the resultant (R) of two orthogonal linearly polarized waves that propagate through a point in the specimen 90° out-of-phase with each other. FIGURE 7.15

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 17 Determination of the resultant wave from its components at individual points in time. FIGURE 7.16

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 18 A single Cartesian coordinate system showing the propagation of a counterclockwise circularly polarized wave. FIGURE 7.17

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 19 A single Cartesian coordinate system showing the propagation of a clockwise circularly polarized wave. FIGURE 7.18

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 20 Determination of the resultant of two orthogonal linearly polarized waves that propagate through a point in the specimen 180° out-of-phase with each other. FIGURE 7.19

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 21 Determination of the resultant wave from its components at individual points in time. FIGURE 7.20

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 22 Linearly polarized light parallel to the azimuth of maximal transmission of the analyzer. FIGURE 7.21

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 23 Determination of the resultant of two orthogonal linearly polarized waves that propagate through a point in the specimen 90° out-of-phase with each other. FIGURE 7.22

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 24 Determination of the resultant of two orthogonal linearly polarized waves that propagate through a point in the specimen 0° out-of-phase with each other and through the azimuth of maximal transmission of the analyzer. FIGURE 7.23

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 25 Linearly polarized light parallel to the azimuth of maximal transmission of the polarizer. FIGURE 7.24

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 26 The intensity and ellipticity of the light passing through a first-order wave plate is wavelength dependent. FIGURE 7.25

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 27 The color of a birefringent object in a polarized light microscope with a first-order wave plate depends in part on the orientation of the fast and slow axes of the specimen relative to the slow axis of the compensator. FIGURE 7.26

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 28 The colors produced by a first-order wave plate, in the absence (A) and presence of a specimen with its slow axis oriented parallel to the slow axis of the compensator (B) and perpendicular to the slow axis of the compensator (C). FIGURE 7.27

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 29 The colors of a specimen composed of positively birefringent molecules is a function of the orientation of the molecules. What colors would we observe in the specimens above if they were composed of negatively birefringent molecules? FIGURE 7.28

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 30 The structure of positively birefringent molecules (e.g., starch and cellulose) and a negatively birefringent molecule (DNA). FIGURE 7.29

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 31 Compensation with a Berek or Ehringhaus compensator. The compensator is tilted from the horizontal position to the vertical position to compensate the specimen. FIGURE 7.30

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 32 The ranges and sensitivies of various compensators used with polarized light microscopes. FIGURE 7.31

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 33 Positively birefringent stress fibers in cheek cells with two different orientations are brought to extinction with a Brace-Köhler compensator. FIGURE 7.32

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 34 A de Sénarmont compensator typically utilizes a λ/4 plate and a rotating analyzer. FIGURE 7.33

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 35 The phase relations between the clockwise and counterclockwise circularly polarized waves that leave the quarter wave plate for a specimen with a phase angle of 45 degrees (A-D) and a specimen with a phase angle of 90 degrees (E-G). When the counterclockwise circularly polarized wave is 45° ahead of the clockwise circularly polarized wave, the resultant linearly polarized wave is rotated 22.5° counterclockwise relative to the axis of maximal transmission of the polarizer. To extinguish the resultant, the analyzer also has to be rotated 22.5° counterclockwise. When the counterclockwise circularly polarized wave is 90° ahead of the clockwise circularly polarized wave, the resultant linearly polarized wave is rotated 45° counterclockwise relative to the axis of maximal transmission of the polarizer. To extinguish the resultant, the analyzer also has to be rotated 45° counter clockwise. FIGURE 7.34

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 36 When the counterclockwise circularly polarized wave is 0° ahead of the clockwise circularly polarized wave, the resultant linearly polarized wave is rotated 0° clockwise relative to the axis of maximal transmission of the polarizer. To extinguish the resultant, the analyzer also has to be rotated 0°. FIGURE 7.35

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 37 The retardation of the mitotic spindle is temperature-dependent. FIGURE 7.36

Companion site for Light and Video Microscopy. by Wayne Copyright © 2009 by Academic Press. All rights reserved. 38 The entropy and enthalpy of polymerization of spindle fibers can be determined by drawing a vant Hoff plot with ln ([B]/[Ao–B]) vs 1/T. FIGURE 7.37