BasicsCoursePhDprog-PeterEvennetOctober2012

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PhD Program: Basics of Light Microscopy - Peter Evennett / LMF - Course October 2012

Setup -Fri 05 October

Setup of all needed course material in Small Auditorium

Equipment needed

  • Microscopes
  • Optical bench
  • Spinning disk gadget
  • Polarazing sheets
  • Diffraction Grids
  • Fluorescent samples
  • Fluorescent Beads (Prepare slides)
  • First day Handout for students
  • Light torches
  • cameras and monitors
  • Cleaning stations for each table
  • condenser screwdrivers for each microsocpe
  • oil/DIC objectives for each microscope (next to stand)
  • 2 polarization filters for each microscope

DAY ONE (8th October)

FROM TO ACTIVITY TOPICS Responsible person(s) Materials needed
9:00 09:30 FINAL PREP Test everything works LMF
9:40 10:25 INTRODUCTION COURSE INTRO
WHO is WHO? Round table
LMF TEAM White board and pen
10:25 11:05 Session 1
Theory
PRINCIPLES OF LIGHT MICROSCOPY
Historical aspects
Very basic components and principles ( incl. Refraction-basics)
PETER EVENNETT (JAN) Jan shortly showing airy pattern from hole in mirror at one of the teaching microsopes (+ screen)
11:05 11:15 BREAK BREAK
11:15 11:50 SESSION 1
Practice
SHORT (5min !!!) Rotation
capillary/plastic (Huisken) in oil demo
Coin-in-cup demo
red and green laser through milky water with black background
(PETE)Jan
PETER
SILKE (Britta)
*capillaries, oil
* coin-in-cup, water
*laser pointer, glass with "milky" water and black background
11:50 12:45 SESSION 1
Theory
Introduction to lenses (Lens demo fun)
Lens aberrations (see colour fringes of lighting image with simple lens)
Resolution, Numerical aperture
Magnification
PETER EVENNETT little lenses
12:45 13:45 LUNCH LUNCH
13:45 14:45 SESSION 1
Practice
15min (!!!) Rotation
Milky Glass Block demo
Show and discuss microscope parts (upright and inverted)
NA measurements
JAN
BRITTA
SILKE
*milky glass block, teaching microscope, coloured filters for microscope, iris objective
*slides with arrows, simple upright and inverted microscope
*horizontal protractor on stand, objective holder, 2 air objective with different (!) NAs (and maybe an iris objective), calculator, pencils, rubber
14:45 15:30 SESSION 2
Theory
Microscope illumination
Conjugate planes
PETER EVENNETT
15:30 15:45 BREAK BREAK
15:50 17:05 SESSION 2
Practice
(30min Rotation)
conjugate planes on Peters microscope
conjugate planes on optical bench
PETER EVENNETT
JAN
*Peters microscope setup
*optical bench demonstrating a transmitted light microscope, incl. light source
17:05 17:25 SESSION 3
Theory
Koehler Illumination
Epi illumination
(Microscope alignment)
PETER EVENNETT
17:15 18:30 SESSION 3
Practice
Koehler illumination
(Microscope alignment)
LMF TEAM students manual microscopes, screwdrivers, nice brightfield sample

DAY TWO (9th October)

FROM TO ACTIVITY TOPICS Responsible person(s) Materials needed
9:00 10:00 Recap day 1 LMF TEAM Questions
10:00 10:45 SESSION 1
Theory & Practice
Eyepieces
Depth of Field
Depth of Focus
PETER EVENNETT
LMF TEAM
students microscopes with 10x/0.25 and 40x/0.65 objectives
Stereoscope with a "sample"
10:45 11:00 BREAK COFFEE BREAK
11:00 12:45 Session 2
Theory & DEMONSTRATION
Diffraction and Diffraction slide fun
Diffraction string show
ABBE'S DIFFRACTION EXPERIMENT
Peter's Video (50min)
PETER EVENNETT Diffraction slides, torch, "dashed" string
12:45 13:45 BREAK LUNCH
13:45 14:15 SESSION 2
Demo and Theory
Dan's diffraction setup demonstration SILKE Dan's ABBE Diffraction demo setup
14:15 15:00 SESSION 3
Theory and Demo
Lens aberrations and corrections PETER EVENNETT
JAN
magnetic lens + laser suitcase on white board|
15:00 15:30 SESSION 4
Theory
Introduction to contrast PETER EVENNETT
15:30 15:45 BREAK COFFEE BREAK
15:45 16:05 SESSION 4a
Theory
Dark Field microscopy PETER EVENNETT
16:05 16:30 SESSION 4a
Practice
Dark Field microscopy LMF TEAM diatome slides
16:30 16:50 SESSION 4b
Theory
Basics of Phase Contrast microscopy PETER EVENNETT
16:50 17:30 SESSION 4b
Practice
Phase Contrast microscopy LMF TEAM cheek cell samples
17:30  : SESSION 4b
Theory
Details of Phase Contrast microscopy PETER EVENNETT

DAY THREE (10th October)

FROM TO ACTIVITY TOPICS Responsible person(s) Materials needed
9:00 10:00 Recap day 2 LMF TEAM Questions
10:00 10:30 SESSION 1
Theory
Objective reading PETER EVENNETT
LMF TEAM
objectives, eyepieces, polylux
10:30 10:45 BREAK BREAK BREAK
10:45 11:45 SESSION 1
Practice
Group rotation
Cover glasses, sample mounting and Objective cleaning
Objective reading
JAN
BRITTA, SILKE
Coverglasses (High preciscion), different oils, slides, different cleaning reagents, cover glass thickness measure meter
at least 7 different, "interesting" objectives (for a group of 6-7)
12:15 12:45 SESSION 2
Theory
Polarized light microscopy PETER EVENNETT
12:45 13:45 BREAK LUNCH
14:00 14:40 SESSION 2
Practice
Polarized light microscopy LMF TEAM Hair, stone samples, benzocain cristals, rulers, gummi bears, plastic dishes, ...
14:40 15:05 SESSION 3
Theory
Differential Interference Contrast (DIC) PETER EVENNETT
15:05 15:40 SESSION 3
Practice
Differential Interference Contrast (DIC) LMF TEAM cheek cell samples
15:40 16:00 BREAK BREAK BREAK
16:00 16:15 Question round Questions to Peter Evennett PETER EVENNETT
LMF TEAM
16:15 17:50 SESSION 3
Theory & Practice
Detectors BRITTA • CCD chip
• example cameras showing different chip sizes
• RGB Slider Spot Camera setup
Demo: CCD chip under stereo microscope
17:50 18:15 SESSION 3
Practice
PIXELS LMF TEAM Sesame seeds, sunflower seeds, nudels, gumibears and a checker board

DAY FOUR (11th October)

FROM TO ACTIVITY TOPICS Responsible person(s) DEMO Activities Materials needed
9:00 10:00 Recap day 3 LMF TEAM
10:00 10:45 SESSION 1
Theory
Fundamental concepts of fluorescence JAN - -
10:45 11:00 BREAK BREAK BREAK BREAK BREAK
11:00 12:15 SESSION 2
Theory & Practice
Fluorescence Microscopy SILKE • Light sources
• Lamp houses
• Spectra
• Filters
• Filter cubes
• Filter spectral charts of each microscope (laminated)
• Spare lamp houses
• Lamps
• Ocean optics equipment
• Four (4) different Filter-sets (cubes), with the appropriate descriptions
• diagramms for drawing filter/fluorophore spectra
12:15 13:00 SESSION 2
Practice
Fluorescence imaging at microscopes LMF TEAM - • Fluorescent labeled samples
13:00 14:00 BREAK LUNCH LUNCH LUNCH LUNCH
14:00 15:45 SESSION 3
Theory & Practice
What is a pixel
Analog to Digital conversion
Nuyquist-Shannon
Quantitative Imaging
Spatial Calibration
BERT
DAVIDE
15:45 16:00 BREAK BREAK
16:00 18:00 SESSION 4
Practice
Imaging with CCD cameras using LMF systems LMF TEAM • Displaying
• Pixel size
• Look up table
• Binning
• Exposure time
• Saturation
• Signal to noise
• ROI
• Bit depth
• Auto map
• Auto contrast
• Microscopes:
DVcore(Davide, 2 students)
N-Storm (Britta, 3 students)
p-DV (Bert, 2students)
L1 (Pete, 2 students)
self-built-syst (Jan, 3 students))
• Fluorescence Sample slides (Kidney / Cells)
19:00 open end Dinner with Peter LMF TEAM

DAY FIVE (12th October)

FROM TO ACTIVITY TOPICS Responsible person(s) Materials needed
9:00 10:30 Recap day 4 LMF TEAM Question sheets (!)
10:30 10:45 BREAK BREAK BREAK note: break might have been too early? (usually in between the Optical sectioning talk)
10:45 13:00 SESSION 1
Theory & Demo
Optical sectioning methods
Pros&Cons
JAN "confocal equipment":
laser pointer, mirror, broken scanning mirror
dual laser pointer (green and red) to demonstrate penetration of diff. wavelength
13:00 14:00 LUNCH LUNCH LUNCH
14:00 15:00 SESSION 2
Small Groups Discussion
What techniques might be good for me
(Own Projects)
LMF TEAM
15:00 15:15 BREAK BREAK BREAK
15:15 16:00 SESSION 3 Course evaluation LMF TEAM
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