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Review from last lectures

Blood-contacting materials
Rationale for studying protein
adsorption
Case Study: Fibrinogen
Motivation
• Implants can often lead to blood clotting
• Design of biomaterials requires a
fundamental understanding of hemostatic
mechanisms and how these impact response
to biomaterials

Tom Horbett (Univ of Washington)


Blood clotting
• Components:
– Platelets; fibrinogen; vWF
• Phases in blood clotting pathway: platelet
activation, adhesion, aggregation, clot
retraction, clot dissolution
• Crosslinking between platelets (via cell surface
receptors) and serum proteins
• Serum proteins can adsorb onto biomaterial
surfaces
Why is fibrinogen adsorption assessment
required for new biomaterials?
Lecture Outline
• Proteins found in blood
• Protein adsorption models
– Langmuir isotherm
– Assumptions of model
– Limitations of model
Properties of the “Big 12” plasma proteins
Motivation for Adsorption Theories
• To reduce experimental findings into a concise
physical description, and eventually, into a
physical law
• A physical law can then predict behavior of
interactions between proteins and other
surfaces
Kinetic Protein Adsorption Models

• transport towards the interface


• attachment at the interface
• eventual structural rearrangements
in the adsorbed state
• detachment from the interface
• transport away from the interface
Definitions
• Adsorption: molecule adheres to the surface
• Absorption: molecules go into the interior of the
surface
• Physical (physisorption): molecule is bound by
physical forces
• Chemical (chemisorption): chemical bond is
formed

*How can you tell the difference between physical


and chemical adsorption?
Desirable outcomes from model:
• Information about adsorption isotherms
• Adsorption kinetics (in situ)
• Conformation of adsorbed proteins
• Number and character of surface bound protein
segment
• Physical parameters describing the adsorbed
protein layer

*How would you validate these predictions


experimentally?
Start with simple system
(proposed by Irving Langmuir in 1916)
• Dependence of surface coverage of adsorbed
inert gas on partial pressure of gas above
surface at fixed temperature (isothermal)
• Known as Langmuir isotherm
• Developed for non-interacting simple gases
• Extensively employed to analyze
macromolecular adsorption processes in
biology
Assumptions of Langmuir model
• All surface sites have same activity for
adsorption
• Adsorbed species do not interact with each
other  only a monolayer can form
• Monolayer is an ideal 2-d solution of equal
sized solute and solvent molecules
• Competition between solute and solvent to
occupy a site on the surface
Derivation: Key Relations

x2  x  x  x1
s
1
s
2
s b
aa
K 2 1
s b
aa1 1
x: mole fraction
a: activity
1: solvent
2: solute
s: surface
b: bulk
Derivation
Assume that surface layer is ideal. Activity for the surface can be
replaced with mole fraction.
s b
xa
K 2 1
s b
xa 1 2
Surface contains only two components: solvent and solute.
b
a
s b K 2
b
xa a
K  x2 
2 1 s 1
(1  x )a2
b s
2 a b
K 2 b 1
a1
Derivation
Define a new constant b = K/a1b
Define θ = fraction of surface occupied by solute
b
ba
x  b
s
2
ba2  1
2

For ideal solutions, a=c


bc kc
 
bc  1 kc  1
k: equilibrium constant for
adsorption
Langmuir Adsorption Isotherm
• Case 1: kc is large (kc>>1)
kc
 – Θ approaches 1, surface is saturated
kc  1 • Case 2: kc is small (kc<<1)
– Θ ~ kc for low c

Surface concentration initially increases linearly with solute


concentration and switches to an asymptotic approach to
monolayer saturation
Alternative Derivation
Start from kinetic equation

d
 ka c(1   )  kd
dt
Set equal to zero (assume equilibrium)

kc

kc  1
Adding Complexity
Models can be made more complex by adding
appropriate changes, eg: two-domain adsorption
Two-Domain Langmuir Isotherm
Model
Start from kinetic equations
d1
 ka1c(1  T )  kd1  ka 21 (1  T )  kd 2
dt
d 2
 2ka 21 (1  T )  2kd 2
dt
Set equal to zero (assume equilibrium)
1 1 1 1 1 1 2 1 1
T   1 (  ) 
2 K1K 2 c 2 K 2 2 K1K 2 c 2 K 2 K1K 2 c
Two-Domain Adsorption

Two-domain protein surface coverage values are higher than one-


domain protein surface coverage values; this difference is most
significant at low concentrations.
Suggested Reading: 1_5 Can et al.
Assigned Reading for 2/3: 1_6 Krishnan et al.