1 Light Rays and Light Waves

Special methods of optical microscopy 1
(Prof. Dr. R. Schreiner)
What is light?
Physics: „Description“ of phenomena in nature by the use of (more or less simplified) models with are in agreement with
observation
 Mathematical description possible
Physical Models for light:
light beams (light rays),

light waves,
light particles (photons)
1. Wave optics (light propagation)
1.1. Light rays
1.2. Light waves

1.1. Light rays (geometical optics) (Prof. Dr. R. Schreiner)
a) viewing angle and magnification

Example: Magnification of optical instruments (resp. Diminuation  optical
micro lithography in semiconductor industry)
The optical path is reversible!
approaching the object
visual angle
s: minimum sharp visible range
Extension of the viewing angle by approaching the

object to the eye.
1.1. Light rays (Prof. Dr. R. Schreiner)

magnification of the ocular (magnifying glass)
virtual image viewing angle:
magnification:
ocular (magnifying glass)
virtual image at b
„normal magnification“
object at s
Problem: large Vok require very small fok  difficult, expensive… solution: additional lens: objective (lens system)

virtual (observed) image: B real intermediate image: Bob=Gok
Object: G angle of sight σ

(without microscope)
objective lens
variable:
movable
move lens to focus until: ocular lens
ocular
objective lens:
lens:
b) Microscope objectives
infinity optics
Ocular Ocular (eyepiece)

intermediate
Image plane
Tube lens
parallel optical path in

the „infinite-space“
Objective
objective
specimen plane
tube lense (e.g. Zeiss: f=164,5 mm)

b) Microscope objectives Single lenses show unavoidable aberrations

Solutions: aspheric lenses, lens systems
blue red
achromatic lens system
achromatic systems
↓
half apochromatic lenses
↓
apochromatic lenses
c) Microscope ocular (Lens System: ocular and objective)

virtual (observed) image: B real intermediate image: Bob=Gok
blue red
Object: G angle of sight σ

(without microscope)
achromatic lens system
objective lens
movable
move lens to focus until: ocular lens
ocular lens (eyepices)

d) field of view
objective
objective
Size of the intermediate image in the ocular BOb (=„field of view number“ S, e.g. S=23 mm
“black edge” of the image)
S
G
VOb
objective 100x → 0.23 mm

ocular 10x → 23 cm
e) Length measurements using eyepiece reticles
Example: objective magnification100x:

object size = 0,7mm/100 = 7µm
Conclusion:
Model „light beams“ (light rays) explain the magnification and construction of optical instruments
but does not explain:

- Optical resolution (it is not possibe to get to extremely high magnifications or diminuations)
- How does a lens work (light refraction)
 Redefined model required…
 Question: What kind of rays do you know?

1.2 Wave optics: resolution and contrast (Prof. Dr. R. Schreiner)
1.2 Wave optics: resolution and contrast (Prof. Dr. R. Schreiner)
The image quality of an optical system depends not on magnification, but on

the optical resolution and the image contrast.
1.2.2. Resolution of an
objective lens
diffraction rings
objective and tube lens
pinhole in object plane
screen 2 closely neighboring

Diameter of the airy disk: objects
top view
side view:

d min 
SIN min
1.2 wave optics: resolution and contrast (Prof. Dr. R. Schreiner)
a) Plane illumination (plane wave)

„screen“
optical path difference
(„optical retardation“)
reflected light waves
tiny object
(reflects light
waves)
parallel illumination
(reflected light mode)
„plane waves“
diameter of the airy disk:

interference of the wavelength
reflected light waves: maximum: resp.

minimum: d min 
SIN min
0,5Dob
the airy disk is limited by 1st. diffraction minimum: with  min  arctan
f ob

d min_ Fl 
n  SIN min
b) Influence of illumination angle on optical resolution

effect of the illumination angle:
1st diffraction
minimum
object
illumination
optical path difference (“optical retardation”)

d min 
 
2  N . A.Objektiv
objective
d min  
SIN min  SIN Objektiv  N . A.illumination
N . A.objective Beleuchtung
resolution

d min  dmin≈0,3 µm (at 550 nm)
2  N . A.Objektiv
objective
human eye (s=25cm): 200 µm
→ Vf=200 µm / 0,3 µm ≈ 700 x-times
„useful magnification“
„blank magnification“:
ways to improve the resolution


d min 
2  N . A.objective
1. decrease the wavelength
► UV
► DUV
h h
► REM  
p me  v
2. near-field optical microscopy (SNOM)
reference: PTB Braunschweig

c) contrast Even if you have the best resolution a object is only visible with sufficient contrast!
I max  I min ► illumination

K ► reflection
I max  I min ► fluorescence
possibilities for increasing the contrast
Improving the contrast by variation of the illumination
bright field
dark field
daylight
e.g. surface of cast iron

bright field dark field
afternoon sun
(Zeiss)
possibilities for increasing the contrast ► illumination

Increasing the contrast by polarization or interference ► reflection
- Polarization contrast
► florescence
analyzer
e.g. cast iron
polarizer bright field polarization
contrast
bright field pol. contrast pol. Contrast and lambda plate
Interference of the wavelength

reflected light waves: maximum:
minimum:
d) possibilities for increasing the contrast ► illumination

Increasing the contrast by polarization or interference ► reflection
Interference contrast (DIC)
► fluorescence
optical path difference:
Interference of the wavelength

reflected light rays: maximum:
minimum:
interference contrast
e.g. „leaf surface“ (biol.)
bright field differential interference contrast
interference contrast („DIC“ of a semiconductor chip surface)

comparison of contrast methods
bright field
dark field
interference contrast (DIC)
1.2. wave optics: “sharpness” of an image (Prof. Dr. R. Schreiner)
sharp
d) „sharpness“ dull
grating
=combination of
resolution and contrast blurred
object curve
physical limit
image curve
modulation-transfer-function
spatial frequency [lines/mm]
contrast-transfer of the Leica Plan:

spatial frequency
object: line grid contrast-transfer-function:
modulation transfer of the line grate
picture: washed out
good system
object modulation
poor system
picture modulation
line pairs/mm
sharpness and contrast: modulation-transfer-function

focus plane
“depth of sharpness” high

depth of sharpness:
resolution limit
“depth of sharpness” small

aperture angle of the
optical system
aperture diaphragm
(in the illumination path) increasing the contrast by closing the aperture diaphragm:
blurred
depth of sharpness:
sharp
object
sharp object borders,

high contrast
focus-plane
depth of sharpness Ts: height range, in that the object details can be
reproduced sharply
increasing the contrast by closing the aperture diaphragm:
(Zeiss)
The luminous field is the projection of the luminous field diaphragm:

illumination (simplified)
Luminous-field diaphragm (“object”)
aperture (iris diaphragm)
beam splitter
object plane
to the field of view:

ocular
luminous field:
(“picture” of the
luminous field aperture)
Luminous-field diaphragm: open closed
luminous field- and aperture-diaphragm

a) Using minor depth of sharpness for thickness measurement
Fine adjustment
coarse adjustment
Focussing drive
Quelle: Zeiss
a) Application of small depth of sharpness for profilometry
confocal microscopes (e.g. Zeiss LSM5):

C. “sharpness” of the microscope picture (Prof. Dr. R. Schreiner)
b) Application of high depth of sharpness for 3D-imaging
(stereomicroscope /„surgical microscope“)
stereomicroscope
References:
Zeiss, Leitz
Conclusion:
The model „Light wave“ is sufficient for the understanding of many aspects of technical optics
Open Questions:
- What kind of wave is it?

- Propagation velocity of the wave?
- How to create this waves and what are the causations?
 Refinded model required
 light as electromagnetic wave (Maxwell Theory)

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1 Light Rays and Light Waves

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Special methods of optical microscopy 1

(Prof. Dr. R. Schreiner)

 Mathematical description possible

Physical Models for light:

light beams (light rays),

1. Wave optics (light propagation)

1.1. Light rays

1.2. Light waves

a) viewing angle and magnification

The optical path is reversible!

approaching the object

s: minimum sharp visible range

Extension of the viewing angle by approaching the

a) viewing angle and magnification

ocular (magnifying glass)

a) viewing angle and magnification

Object: G angle of sight σ

move lens to focus until: ocular lens

Ocular Ocular (eyepiece)

parallel optical path in

tube lense (e.g. Zeiss: f=164,5 mm)

b) Microscope objectives Single lenses show unavoidable aberrations

achromatic lens system

c) Microscope ocular (Lens System: ocular and objective)

Object: G angle of sight σ

ocular lens (eyepices)

objective 100x → 0.23 mm

e) Length measurements using eyepiece reticles

Example: objective magnification100x:

but does not explain:

 Redefined model required…

 Question: What kind of rays do you know?

The image quality of an optical system depends not on magnification, but on

objective and tube lens

pinhole in object plane

screen 2 closely neighboring

a) Plane illumination (plane wave)

diameter of the airy disk:

b) Influence of illumination angle on optical resolution

optical path difference (“optical retardation”)

human eye (s=25cm): 200 µm

→ Vf=200 µm / 0,3 µm ≈ 700 x-times

ways to improve the resolution

reference: PTB Braunschweig

I max  I min ► illumination

possibilities for increasing the contrast

Improving the contrast by variation of the illumination

e.g. surface of cast iron

possibilities for increasing the contrast ► illumination

bright field pol. contrast pol. Contrast and lambda plate

Interference of the wavelength

d) possibilities for increasing the contrast ► illumination

optical path difference:

Interference of the wavelength

e.g. „leaf surface“ (biol.)

bright field differential interference contrast

interference contrast („DIC“ of a semiconductor chip surface)

comparison of contrast methods

spatial frequency [lines/mm]

contrast-transfer of the Leica Plan:

picture: washed out

sharpness and contrast: modulation-transfer-function

“depth of sharpness” high