BME 101 Biomedical Optics
and Lasers
Instructor: Irene Georgakoudi
TA: Cherry Greiner
Class meeting time: T/Th: 10:30 -11:45 AM
Office hours: Tuesdays, 3:00-4:30 PM
Blackboard site: http://blackboard.tufts.edu
Reading material on reserve (Tisch):
Handbook of Biomedical Photonics, Tuan Vo Dinh
Introduction to Biophotonics, Prasad
Biological spectroscopy, Campbell
Electo-optics library: Optics, Hecht
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Evaluation
10% Class participation
30% Homeworks/Lab reports (due every Tuesday)
15% Midterm exam
25% Final exam
20% Final paper and presentation
– 10-15 page paper on specific molecular imaging method and
its impact on a clinical problem
» Introduction
» Theoretical background of method
» Background on clinical problem
» Instrumentation
» Methods/Results
» Advantages/Disadvantages
» Suggested improvements
What is biomedical optics?
• Biomedical optics is typically defined as
the area of study of methods/technologies
based on the use of visible light
(applications cover UV-NIR) for:
– Improving basic understanding of biological
processes (from gene to tissue level)
– Enhancing the detection and treatment of
human diseases (from acne to atherosclerosis
and cancer)
Syllabus
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Basic principles
Spectroscopic methods
Microscopy and Imaging
Photodynamic Therapy/Flow Cytometry
Basic Principles
• Light matter interactions


z, t   o cost  kz
– Basic wave definitions
– Schrodinger’s equation
-Bonds and orbitals
-Biological chromophores
Basic Principles
• Laser basics
– Principles of operation
• Stimulated emission
• Critical inversion
• Pumping schemes
– Major laser components
– Laser beam properties
– Diode lasers
Cell and Tissue basics
• Cell basics
– Major cellular components
– Origins of intrinsic cellular
optical signals
• Tissue basics
– Major epithelial types
– Connections to disease and optical
sources of contrast
Spectroscopic methods
• Absorption
• Scattering
• Fluorescence
Epithelium
Connective Tissue
Spectroscopic methods
• Basic Theoretical principles





1
 (r , t )  D 2  (r , t )   a  (r , t )  S o (r , t )  3D  S1 (r , t )
c t
• Instrumentation
• Applications
1
1.5
0
0.5
1
mm
Adenoma
14
12
40-50
30-40
20-30
10-20
1.5
Non-dysplastic
mucosa
10
Normal
8
6
4
2
0
0 2 4 6 8 10
y (cm)
0.5
Intensity (a.u.)
Enlarged
Nuclei, %
Data
Fit
Residual
16
x (cm)
mm
1
18
Adenoma
0.5
atherosclerosis
20
Colon cancer
0
Breast cancer
0
-0.5
800
1000
1200
1400
Raman Shift (cm-1)
1600
1800
Microscopy and Imaging
• Basic Principles
H&E
Confocal in vivo
“En face”
SECTION of
human skin
4-Pi microscopy
Mitochondrial network
of live bacterial cell
80 nm res
Tumors and
blood vessels
imaged in vivo
Triple stained endothelial
Cell of pulmonary artery
Engineered tissue:
Fibroblast (red) in
collagen matrix (green)
Endogenous signal
Optical imaging of cell-matrix
interactions
Collagen gel embedded with GFP-expressing fibroblasts
Leica TCS SP2 confocal microscope
Excitation wavelength: 488 nm
Green channel: 485-490 nm (scattering)
Red channel: 500-620 nm (GFP fluorescence)
Stack size: 238x238x125 m
Images acquired using 63X, 1.2 NA, water immersion objective
Photodynamic Therapy
• Basic Principles
• Applications
Macular
degeneration
Flow Cytometry
• Basic Instrumentations
• Advanced Methods
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Reading assignment:
Introduction to biophotonics, ch. 1 and 2.1
Lecture notes
Also posted on the blackboard site:
– Calculus review
– Electromagnetic waves review
Biomedical optics
Exploiting interactions of light with matter
Wavelengths used typically: 300-900 nm
Why biomedical optics?
• Major advantages: non-invasive; high resolution
• Continuous or repetitive monitoring
• Study/characterize process/disease in natural
environment (no artifacts)
• More sensitive/accurate monitoring
• Real-time information
– Triage with therapy
– Accurate dosimetry
– Psychological impact
A bit of optics history
It all started with the Greeks…
Plato (427-347 BC)
Believer of extramission
theory: Eye emits a
“fire” providing man
the capability of vision
by seizing objects
It all started with the Greeks…
Aristotle (384-322 BC)
Light emitted by a source
is captured by the eyes
when reflected by an
object
It all started with the Greeks…
• Euclid (circa 325-265 BC)
• Treatise entitled
“Catoptrics”
• Foundations of geometric
optics
• First law of reflection
It all started with the Greeks…
Galen of Pergamum
(Claudius Gelenus: 130201 AD)
• Described anatomical
details of the eye
• Identified lens as principle
eye instrument
• Believed in extramission
theory
Philosophers from the Middle-East followed…
Mohammad ibn Zakariya alRazi (864-930AD)
• Also known as Rhazes
• Observed that pupil
contracts in response to
light
Philosophers from the Middle-East followed…
Abu Ali al-Hasan ibn al-Haytham (9651040 AD)
• Also known as Alhazen
• Considered by some as the father of
Optics
• Wrote comprehensive treatise on
optics (Katib-al-Manazir/Book on
Optics), translated in Latin in 1270
– Proved that extramission theory is not
correct
– Detailed description of human eye
– Theory of vision which prevailed until
17th century
– Discussed primary and secondary light
sources, light propagation and colors
– Studied spherical and parabolic
mirrors
– Laws of reflection and refraction
Western philosophers/scientists
Leonardo da Vinci (14521519 AD)
• Initially believed in
extramission, but later
changed his view in
support of external light
sources based on
experiments he performed
with ox eyes
Western philosophers/scientists
Johannes Kepler (15711630)
• Established retinal image
formation theory based on
experiments with ox eyes
• Law of refraction for small
angles of incidence
Theories on nature of light:
Light as a particle vs. Light as a wave
• Only corpuscular
theory of light
prevalent until 1660
• Francesco Maria
Grimaldi (Bologna)
described diffraction
in 1660
Light as a particle
Sir Isaac Newton (1642-1727)
• Embraces corpuscular theory of
light because of inability to
explain rectilinear propagation in
terms of waves
• Demonstrates that white light is
mixture of a range of
independent colors
• Different colors excite ether into
characteristic vibrations--sensation of red corresponds to
longer ether vibration
Light as a wave
Christiaan Huygens (1629-1695)
Huygens’ principle (Traite de la
Lumière, 1678):
Every point on a primary
wavefront serves as the source
of secondary spherical
wavelets, such that the primary
wavefront at some later time is
the envelope of these wavelets.
Wavelets advance with speed
and frequency of primary wave
at each point in space
http://id.mind.net/~zona/mstm/physics/waves/propagation/huygens1.html
Light as a wave
Thomas Young (17731829)
1801-1803: double slit
experiment, showing
interference by light
from a single source
passing though two
thin closely spaced
slits projected on a
screen far away from
the slits
http://vsg.quasihome.com/interfer.htm
Light as a wave
Augustine Fresnel (17881827)
1818: Developed
mathematical wave theory
combining concepts from
Huygens’ wave propagation
and wave interference to
describe diffraction effects
from slits and small
apertures
Electromagnetic wave nature of light
• Michael Faraday (17911865)
• 1845: demonstrated
electromagnetic nature of
light by showing that you
can change the polarization
direction of light using a
strong magnetic field
Electromagnetic theory
• James Clerk Maxwell
(1831-1879)
• 1873: Theory for
electromagnetic wave
propagation
• Light is an electromagnetic
disturbance in the form of
waves propagated through
the ether
Quantum mechanics
Max Planck
• 1900: Max Planck postulates that
oscillating electric system imparts its
energy to the EM field in quanta
• 1905: Einstein-photoelectric effect
Heisenberg
– Light consists of individual energy quanta, photons,
that interact with electrons like particle
Niels Bohr
• 1900-1930 it becomes obvious that concepts
of wave and particle must merge in
submicroscopic domain
• Photons, protons, electrons, neutrons have
both particle and wave manifestations
– Particle with momentum p has associated
wavelength given by p=h/l
• QM treats the manner in which light is
absorbed and emitted by atoms
Louis de Broglie
Schrödinger
Wave definitions
Classical
Description of Light

2

1  E
2
 E 2 2
Wave Equation (derived from Maxwell’s equations)
c t
Any function that satisfies this eqn is a wave

2

1  B It describes light propagation in free space and in time
2
 B 2 2
c t
where,
c  speed of light

E  electric field

B  magnetic induction field
  Laplacian operator
2
(see calculus review handout)
Classical Description of Light
Plane Wave Solution
One useful solution is for plane wave
E
  i kr  i t   i kr t 
E  Eo e e
 Eo e
  i kr t 
B  Bo e
r
where,
k  wave number or propagation vector
B
  angular frequency
Classical description of light
Considering only the real part of the previous
solution to make things simpler, we have for the
electric field propagating along one dimension, z


z, t   o cost  kz

0
(or distance)
Period
time
l  wavelength (m eters)
  period (sec onds)

1

c
frequency (cycles / s or Hz)
 l
2
k
 wavenum ber(convertsdistance to angle)
l
2
  2 
 angular frequency (convertstime to angle)

Light as a wave: Basic concepts
Phase=f=t-kz
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Phase of a wave is the offset of
the wave from a reference point fo
We typically talk about a phase
shift
When light interacts with matter
(e.g. as it travels through a
biological specimen), its speed of
propagation slows down. The
wave emanating from the
specimen exhibits a phase shift
when compared with the initial
wave
The refractive index , n, and the
thickness of a specimen
determine by how much the wave
is retarded
n
c

, where c  speed of light in vacuum
Green-incident wave
Blue-wave after passing
through specimen shifted by
l/4
Phase=f=t-kz
Coherent Light
Monochromatic (only one
wavelength/frequency)
waves traveling in phase
Incoherent Light
Incoherent Light
Monochromatic (only one
wavelength/frequency)
waves traveling out of phase
Constructive
interference
Destructive
interference