1. LESSON 5

2. LLight Microscopy BBright-field microscopy PPhase Contrast microscopy DDifferential Interference Contrast (or Nomarski) DDark-field microscopy FFluorescence microscopy CConfocal Scanning microscopy EElectron Microscopy TTransmission electron microscope (TEM) SScanning electron microscope (SEM)

3. 1. Junqueira’s Basic Histology 1. Junqueira’s Basic Histology ; pp 7 & 9 2. Looking at the Structure of Cells in the Microscope http://www.ncbi.nlm.nih.gov/books/NBK26880/#A1749 ▪ The Electron Microscope Resolves the Fine Structure of the Cell. ▪ Images of Surfaces Can Be Obtained by Scanning Electron Micros copy ▪ Metal Shadowing Allows Surface Features to Be Examined at Hig h Resolution by Transmission Electron Microscopy ▪ Freeze-Fracture and Freeze-Etch Electron Microscopy Provide Vi ews of Surfaces Inside the Cell

5. Resolution, resolution!!!

6. ELECTRONS AND RESOLUTION LIGHT MICROGRAPH ELECTRON MICROGRAPH

7.  Why electron beams?  The wavelength, much shorter than of light  Types  Transmission electron microscope (TEM)  Scanning electron microscope (SEM ) - surface contours TEM SEM R = 0.61λ/ N.A.

10. Electromagnetic lenses a coil of wire through which current flows magnetic field in the center. electrons traveling in their path down the central hole.

11. TEM

12. CONTRAST IN TRANSMISSION ELECTRON MICROSCOPY Electron Transparent Electron Transluscent Electron Dense Dark (Electron dense) (Electron transparent) (Electron transparent) Bright (Electron lucent)

13. Bright (Electron lucent) Bright (Electron lucent) - electrons readily passed Dark (Electron dense) Dark (Electron dense) - electrons absorbed

14. Transmission electron microscope (TEM)

15. SEMTEMDIC

17. SEM

18. Depth of Focus with the Scanning Electron Microscope Light Micrograph of Ant’s Head Scanning EM of Ant’s Head

19. Scanning electron microscope (SEM )

20. Metal Shadowing high resolution three-dimensional replica of the specimen heavy metal shadowing of bacteria to demonstrate flagella

22. Freezing Techniques Freeze-Fracture Freeze-Fracture Freeze-Etch Freeze-Etch

24. TEM Section and Freeze Fracture Comparason Section Through Apical Region ofIntestinal Adsorptive CellFreeze-Fracture Replica of ApicalRegion of Intestinal Adsorptive Cell Tight Junctions TEM Section and Freeze Fracture Comparason Section Through Apical Region of Intestinal Adsorptive Cell Freeze-Fracture Replica of Apical Region of Intestinal Adsorptive Cell Tight Junctions TEM Section and Freeze Fracture Comparason Section Through Apical Region of Intestinal Adsorptive Cell Freeze-Fracture Replica of Apical Region of Intestinal Adsorptive Cell Tight Junctions TEM Section and Freeze Fracture