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Techniques in Light Microscopy

F2020/ PMB 185
Zoom Lectures
MWF 9-10AM

Lab F 1-4


3 Units

Steven Ruzin Ph.D.

Denise Schichnes Ph.D.

Biological Imaging Facility
381 Koshland Hall

The course will be a detailed overview of the practice of light microscopy as applied to scientific investigation. The emphasis of the course will be on the use of the light microscope by biological scientists, however students of other disciplines are welcome. We will cover optical microscope theory, microscope components and mechanics, and all modern optical techniques from Brightfield through Deconvolution microscopy, Structured Illumination, and others. Instruction will consist of lectures and Demonstrations, all virtually via Zoom.

Section 101 (CCN TBA Friday 1–4PM via Zoom

Grading: There will be a Midterm written exam and a written Final Exam. For labs, students will be required to complete a worksheet for each of the labs. In addition, a project is required for the class. It consists of a written paper describing your Virtual forensic analysis of an unknown substance.

Office hour:
Monday 2–3PM,
381 Koshland Hall
Date

Lecture/Lab topic, Fall 2020

Lecture

Topic in Lecture

Lab topic (Fridays)
Aug 26

Introduction

28
Light and Refraction
No Lab
31
Microscope Optics: Lenses annd Conjugate planes
Sept 2
Microscope optics: Imaging and Resolution
4

Lens aberrations (spherical, chromatic),

1: Microscope optics
7
No Classes
9
Microscope configurations: The compound microscope , Microscope Standard Lengths
11

The Condenser: history, optics & Köhler illumination

2: Köhler illumination
Focus Examples
14

Objectives, objective designations, Eyepieces

16

Numerical Aperture and Resolution

18

Darkfield microscopy, Rheinberg Illumination, Oblique Illumination

3: Darkfield
21
Polarized light, Anisotropy, Types of birefringence, Polarized light microscope
23

Polarized light microscopy, Generation of colors,

25

Polarized light microscopy: Quantitative PLM

4: Polarized Light Microscopy
28
Phase Contrast microscopy, presentation, examples, Zernike's Nobel Speech
30

Phase Contrast microscopy

Oct 2
Inverted Microscope, Stereo Microscope,
5: Phase Contrast
5

Interferometry, Differential Interference Contrast Presentation; DIC microscope

7

PlasDIC

9

Hoffman Modulation Contrast

6: DIC
12
Review of transmitted light techniques, Key to determining imaging method
14
Microscope Jeopardy
16

Midterm Exam

7: Microscope Forensics
19

Fluorescence Microscopy, Stokes Shift, Types of sample fluorescence

21

Fluorescence Microscope, presentation

23

Digital Imaging, Nyquist sampling, Digital imaging for microscopy

8: Epifluorescence microscopy
26
Laser scanning confocal microscopy I, presentation
28

Laser scanning confocal microscopy II, resolution

30

Laser scanning confocal microscopy III

9: Laser ScanningConfocal microscopy
Nov 2
Two photon microscopy and resolution, presentation
4

Two photon microscopy II

6

Spinning Disk & Sweep Scanners, Nipkow disks

10: Spinning Disk Confocal Microscopy
9
Deconvolution microscopy, Basis, PSF, Methods,
11

No Class

13
Deconvolution II, Here's a short tutorial on Filtering
11: Deconvolution using HuygensPro
16

Super Resolution: Structured Illumination Microscopy (SIM)

18
Super Resolution: ; PALM/STORM
20
STED
12: Super-Resolution (SIM)
23
Advanced fluorescence imaging: FRAP, FRET, FLIM
25

Holiday: No Class

27

Holiday: No Class

Holiday: No Lab
30

Total Internal Reflection microscopy, TIRF microscope, Implications;

Dec 2
Final Review; Finals Review Resources; Finals Jeopardy; BF Review, Fluor review
4

The History of the Microscope

13: Virtual tour of the Golub Collection
7

RRR Week: No Lecture

9

RRR Week: No Lecture.

11

RRR Week: No Lecture. Project Due 5PM. Upload to bCourses

17

Scheduled 12/17 7-10PM, however, the Final Exam will be a take-home exam. It will be available for download on Dec 14 at 5PM. You'll have 3 days to complete it. Due 12/17 10PM.

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Updated Fri, Nov 27, 2020