Test 2 Flashcards
1
Q
what is adsorption
A
adhesion of molecules to solid surface (no penetration)
2
Q
what affects adsorption the most
A
hydrophobicity and surface charge
3
Q
protein adsorption increases with increasing __
A
hydrophobicity
4
Q
besides hydrophobicity and surface charge, __ can also promote protein adsorption becuause __
A
surface roughness can promote adsorption because proteins will become physically trapped in the valleys of the surface
5
Q
what are the chemical and physical properties governing protein adsorption
A
chemcial
Hydrophobicity
Surface Charge
Physical
Surface Roughness
Steric hinderances
6
Q
rougher surfaces will have a ___ protein adsorption
A
more protein adsorption
7
Q
besides making a surface smoother how do you physically decrease protein adsorption
A
add steric hinderances
8
Q
how do you decrease protein adsorption using steric hinderances
A
add hydrophilic, highly flexible chains on the surface that will physically block the proteins from coming into contact with the surface
9
Q
for steric hinderance for protein adsorption, the chains must be __ because __
A
the chains must be hydrophilic so they can interact with the aqueous environment
10
Q
what is PEG
A
PEG is a large, highly flexible hydrophilic polymer that decreases protein adsorption
11
Q
PEG __ __ protein binding
A
decreases non specific protein binding
12
Q
how does PEG decrease non specific protein binding
A
a large volume of the surface is taken up by these bulky chains that are in constant motion. This prevents protein adsorption
13
Q
PEG is an example of ___ surface property that prevent protein adsorption
A
steric hinderance
14
Q
what are the 4 ideal surface modification techniques
A
(TRDS)
- Thin (to minimize effects on bulk properties)
- Resistant to delamination (want it to stay on the surface)
- Simple and robust
- discourage surface rearrangement after treatment
15
Q
what are examples of covalent surface coating
A
Plasma treatment and self assembling monolayers (SAM)
16
Q
what is plasma treatment
A
assembly of species in a molecularly dissociated gaseous environment
17
Q
what are the possible reactions of plasma treatment
A
- Etching - remove stuff from the surface
- Deposition - add stuff to the surface
- Functionaliztion - covalently add molecules
18
Q
what is the name of the advantages of plasma treatment
A
Ratner and Hoffman
19
Q
What the the advantages of plasma treatment
A
(g clefs)
- conformal - plasma will follow shape of implant
- free of voids
- easily prepared
- good adhesion to surface
- sterile
- low leachable substances
20
Q
what are disadvantages of plasma treatment
A
- hard to predict composition of surface that will result
- expensive
- difficult to form uniform reaction
- easily contaminated
21
Q
how are SAMs different than steric hinderance
A
SAMs have hydrophobic regions which will stabilize the structure (steric has hydrophilic chains)
22
Q
why do SAMs attach to the surface
A
it is thermodynamically favored for them to attach and align
23
Q
because they have a hydrophobic region, SAMs are __
A
amphiphilic
24
Q
what are the 3 groups in SAMs
A
- Attachment group
- Long hydrocarbon chain
- Functional Group - can be used to alter hydrophobicity
25
SAMs result in __ reactions
exothermic
26
what is the purpose of the hydrophobic regions in SAMs
they stabilize the structure
27
what are advantages of SAMs
1. ease of formation - can control specific areas where proteins will bind
2. Chemically stable
3. variety of functional groups are possible
28
disadvantages of SAMs
need for particular chemistry to drive attachment
29
what is an example of a non-covalent surface coating
surface modifying agents (SMA)
30
what are SMA
molecules that will spontaneously rise to the surface from the bulk, thus producing a coating
31
Unlike, plasma treatment and SAMs, SMA is a __
pre-fabrication design
32
what allows SMAs to form
there is a driving force to reduce the free energy at the surface
33
Which material is less likely to form SMAs and why
Ceramics because there is less atomic motion in the bulk due to electroneutrality
34
how do SMAs form in polymers
hydrophilic particles inside the polymer will want to move to the surface when the material is placed inside an aqueous environment
35
what determine the effectiveness of SMAs
1. difference in free energy of the surface with and without the SMA
2. Molecular motility of the SMA in the bulk to actually allow it to move to the surface
3. Environment surrounding the implant must be hydrophilic to allow the hydrophilic SMA to rise to the surface
36
what do conversion coatings do (physiochemcial surface modification)
they form a passive layer on metals
37
Conversion coatings can spontaneously form due to __
the porbaix diagram
38
what is a major concern with biological surface modification techniques
maintaining the bioactivity of the biological molecules
39
PEG is an example of a __ for biological coatings
a spacer arm
40
in covalent biological coatings, a molecule can be bound to a __
spacer arm
41
how does a spacer arm work for biological coatings
the spacer arm provides space between the biomolecule and the surface. this allows for greater rotational freedom and improves the activity of the biomolecule
42
a spacer arm is an example of a __ biological coating
covalent
43
give an example of a noncovalent biological coating
a positively charged surface can attract heparin (which is negatively charged)
44
what information does contact angle analysis provide
information about the hydrophobicity of the surface
45
higher contact angel = __
more hydrophobic
46
what is hysteresis (for contact angle analysis)
surface tension of a material can be changed before and after water is added
47
contact angle analysis doesn't provide any info about __
chemical composition of the surface
48
explain how contact angle analysis would appear with SMAs
SMA that would move surface in body is hydrophilic – surface becomes more hydrophilic with time – contact angle decreases with time, so advancing angle is larger
49
what technique would you use to view the 3D surface of a material
atomic force microscopy (ATF)
50
ATF doesn't provide info about __
chemical composition
51
what technique would you use to find the chemical composition of a surface
ATR-FTIR
52
protein binding on hydrophilic surfaces are __
relatively reversible
53
protein binding on hydrophobic surfaces are __
irreversible (protein denature on surface)
54
at early time points, adsorption is ___ controled
diffusion
55
anything that lowers __ is favorable
∆G
56
what is the gibbs free energy equation for protein adsorption
∆Gadsorption = ∆Gprotein +∆Gsolvent + ∆Gsurface
57
adsorption will be thermodynamically favorable if __
the adsorption event lowers ∆G for one or more components without raising ∆G for another component
58
what factors have the largest impact on protein adsorption
(drs)
1. dehydration of the surface and protein
2. redistribution of charged groups
3. structural rearrangment of proteins
59
explain how dehydration of the protein and surface affects protein adsorption
water molecules become more ordered at hydrophobic surfaces
if hydrophobic areas are able to come together through adsorption, they will reduce the hydrophobic surface area that is able to come into contact with water
this will allow water to become more disordered (increase in entropy/disorder is favored)
hydrophobic amino acids will come together to minimize its interactions with water
60
explain how redistribution of charge affects protein adsorptoin
adsorption is preferred when the material and the surface have opposite charges
in this case, the transfer of fewest ions is required
repulsion of material and surface can be overcome by adsorption of ions from the solution to help create opposite charges
but the adsorption of ions costs energy
61
explain how structural rearrangement of the protein affects protein adsorption
a less stable protein will adsorb more easily since conformational rearrangement is easier
allow for optimal conformation so that hydrophobic and charged regions of the protein can be placed to fulfill the two other properties (dehydration and redistribution of charged)
62
amino acids are considered __ because they can act as either ___
zwitterion - they can act as either acids or bases
63
what are the two amino acids that play a large role in determining 2º, 3º, and 4º structure
proline and cysteine
they are both non charged
64
what about proline and cysteine makes them important for determining 2º, 3º, and 4º structure
proline has a ring backbone that can limit protein folding in 3D
cysteine contains sulfide (SH) which can form disulfide bonds with other cysteine residues
65
__ is the linear order of amino acids
primary structure
66
__ is a disease in the
| primary structure which dictates quaternary structure
sickle cell
67
__ and __ are examples of secondary structures
alpha helix and beta pleated sheets
68
__ are localized interactions between amino acid residues
secondary structure
69
what is the the 3D arrangement of an entire polypeptide chain
tertiary structure
amino acids on the same chain are interacting
70
__ is an example of tertiary structure
myoglobin
71
__ is the primary driving force for tertiary structure formation
hydrophobic interactions between non polar amino acids
minimize the amount of hydrophobic area that is interacting with the water
this is more energetically favorable (entropy)
72
__ is the 3D arrangement of protein subunits
quaternary structure
interactions of amino acids between chains
73
__ is an example of quaternary structure
hemoglobin
74
there is a __ correlating to diffusion
high initial adsorption rate
75
__ is reversible because the proteins are __
initial binding is reversible because proteins are not strongly bound to the surface
76
after __, binding is irreversible
conformational changes
77
why does a plateau occur in protein adsorption kinetics
a plateau occurs due to a monolayer coverage of the entire surface by proteins
78
explain desorption and exchange
a protein with higher affinity for the surface will displace the protein already on the surface
79
why is desorption unlikely after conformational changes occur in the protein
you would have to break all contacts between the surface and the protein
80
what are the techniques for determining protein type and amount
HPLC and ELISA
81
HPLC is used to determine __
the amount of protein in a mixture
82
describe the different types of HPLC
(AANR)
adsorption chromatography - separation by adsorption based on hydrophobic and polar interactions. the first species to exit the column has least affinity for the column
normal phase chromatography - uses a polar stationary phase and a nonpolar mobile phase. more hydrophilic are eluted last since they have affinity for the polar column
reverse phase - hydrophobics elute last
affinity chromatography - integrate area under curve to find out how much of that species is present
83
describe the process of an ELISA
1. primary antibodies are added first
2. the sample is then added
3. the secondary antibody is added that will bind to the protein. the secondary antibody has an enzyme attached to it
3. a substrate for the enzyme is added
color change is proportional to the amount of protein present
84
what does a modified ELISA tell?
used to determine the amount of protein already on the surface of a material
85
how is modified ELISA different from an ELISA
in a modified ELISA, the protein is already attached to the surface. therefore you don't need to used the primary antibodies normally used in an ELISA
86
a negative result in an ELISA can mean__
there is no protein or the protein is denatured
87
__ have attained tissue -specific functions
differentiated cells
88
__ can differentiate into a variety of cell types
progenitor cells
89
what are proliferation assays?
that are basically viability assays over time
90
when cell spreading, __ interacts with __ to anchor the cell firmly in place
the integrin interacts with the ligands
91
describe the process of cell migration
1. pseudopodia extend
2. membrane attaches to the substrate via integrin
3. after pseudopodia are adhered, there is a contractile force and the release of rear receptors
4. integrin receptors are recycled from back to front
92
__ is the speed of cell movement
translocation speed
93
__ is the time the cell moves without changing direction
persistance time
94
what are the cytotoxicity assays
Direct contact assays
Agar diffusion assay
elution assay
95
in a __, the material is placed over top of the cells and cells death in the surrounding area is observed
direct contact assay
96
in a direct contact assay, you need __
both a positive (non-cytotoxic) and negative (cytotoxic) control
97
what is an agar diffusion assay
cells are covered with agar before the material is placed on top
soluble products from material diffuse into the agar
98
describe the results seen from an agar diffusion assay
if cell death occurs farther away from the sample, the more cytotoxic the sample
99
__ are performed to determine the cytotoxicity of leachable molecules found in biomaterials
elution assays
100
describe how a adhesion/spreading assay works
1. allow cells to adhere to surface then wash away non-adhered cells
2. number of cells -on suface = total number of cells - cells washed away
101
what are the disadvantages of cell adhesion/spreading assays
concerns over reproducibility of the force exerted in the washing step
102
what are the migration assays
capillary tube test
boyden chamber assay
103
describe a capillary tube test
1. confine cell population in a small tube.
2. the opening of a tube is placed against a surface, allowing the cells to migrate out in a fan like shape
3. the area of the fan shape is measured
104
disadvantages of capillary tube test
it is difficult to distinguish the effects of proliferation and migration
105
what does a boyden chamber assay tell
looks at how cells might migrate in response to materials that are leached
(cells might move closer to the leached material)
106
this assay is used to determine the effects of soluble factors on migration rather than examining the impact of the substrate
boyden chamber assay
107
how does a boyden chamber assay work
1. filter separates the cells from the soluble agents
2. cells are seeded on the top layer and the substance is on the bottom layer
3. cells will move to the bottom chamber if they are attracted to the leached materials
108
how do you track individual cells
1. put cells on material
2. take pictures to determine where they move over time
speed of migration
persistence time
109
speed of migration and persistence time are associated with __
tracking individual cells
110
longer persistence time =
chemotactic response
111
what type of immunity is always present
innate immunity
112
__ provides the first line of defense
innate immunity
113
if the invading organism is not cleared by the innate immune response, then the body engages the _
acquired immune response
114
what are the components of the innate immune response
1. barriers (skin)
2. physiologic barriers (pH in stomach)
3. Phagocytic cells
4. inflammation
115
what are the two types of inflammation
acute
| chronic
116
__ occurs immediately after an injury
acute inflammation
117
of the two types of inflammation, which one is undesired
chronic inflammation
118
__ inflammation persists over weeks to months
chronic inflammation
119
redness, swelling, heating, pain are signs of __
acute inflammation
120
Acquired immunity involves the activation of __
white blood cells called lymphocytes
121
acquired immunity responses to a particular _
antigen
122
when does infection develop2
if the invader persists after exposure to both the innate and acquired immune response
123
red blood cells are known as
erythrocytes
124
lymphocytes are part of the __
acquired immune response
125
granulocytes are __
phagocytic cells in innate immunity
126
neutrophils are a type of ___
granulocytes
127
what is the first step in the timeline of an injury (acute inflammation)
tissue macrophages release cytokines to call in neutrophils (chemotaxis)
128
after tissue macrophages call neutrophils, what happens in the timeline for acute inflammation
neutrophils arrive and move out of the blood and into the tissue via extravasation
129
describe the process for extravasation
1. rolling - selectins are upregulated in inflamed endothelium. the selectins interact with the neutrophils which helps bind them to the endothelium
2. activation - neutrophils are activated by cytokines that interact with integrins on the neutrophils. once activated, the integrins have higher affinity for CAMs on the endothelium
3. Arrest and adhesion - strong interactions between integrin on neutrophils and CAM on endothelium cause the neutrophil to adhere to the endothelium
4. Migration - diapedesis, the neutrophils squeezes through the endothelium cells and into the tissue
130
after extravasation, what do the neutrophils do in acute inflammation
once in the itssue, the neutrophils will phagocytose
they can undergo respiratory burst - create reactive oxygen species to kill invaders
they secrete cytokines that recruit monocytes
they can secrete cytokine that release factors that are chemotactic for lymphocytes
***provides a point of communication or crossover between the innate and acquired immune system ***
131
after the neutrophils secrete cytokines to recruit monocytes, what happens
monocytes arrive and become macrophages
132
how do macrophages work
phagocytosis
respiratory burst
**secrete cytokines to increase acquired immune response**
**act as antigen presenting cells***
133
describe a potential problem during phagocytosis by macrophages
small particles - macrophages will eat non degradable particles. but the particle will be re-released when the macrophage dies (silicosis)
large particles - large particles undergo frustrated phagocytosis. the macrophage will release lysosomal enzyme outside of the cell to digest the invader. too much of this can lead to tissue damage
134
how to terminate acute inflammation
short half life of chemical mediators
cytokines are produced that can act as inhibitors to pro-inflammatory cytokines
135
what are the types of cells that can be used for assaying acute immune response
leukocytes
endothelial cells
136
leukocyte cell assays are used for assaying what type of cells
neutrophils and macrophages
137
what can you determine using leukocytes cells in an assay
adhesion/spreading - more spreading means more inflammatory
cell migration - more migration means more inflammatory (boyden chamber assay)
pro inflammatory cytokine release - use ELISA to detect pro inflammatory cytokines
look at cell surface markers - use FACS (similar to ELISA but works on cell surface) increase in markers/ receptors means more inflammatory
138
what can you determine using endothelial cell in an assay to
increase in cell surface receptors on endothelial cells (selectin, CAMs) means more inflammation
This promotes the migration of neutrophils
139
1-3 days after the injury, macrophages call in __ to __
call in fibroblasts and vascular endothelial cells
140
proliferation of fibroblasts and vascular endothelial cells leads to the formation of __
granulation tissue
141
granulation tissue may form part of the __
foreign body reaction
142
what are foreign body giant cells (FBGCs)
they are fused macrophages which form in an attempt to phagocytose the biomaterial, which is much larger than a single cell
143
why does foreign body giant cells form
to attempt to phagocytose biomaterials which are much larger than a single macrophage
144
what type for foreign body reaction will the following materials get:
smooth implants
rough implants
high surface area to volume (porous)
Low surface area to volume
smooth - get macrophages
rough - get macrophages and FBGCs
high SA - marcophages and FBGCs
low SA - fibrous (granulation tissue)
granulation tissue has more collagen and has higher mechanical properties
145
how long do FBGCs remain
they may remain for the lifetime of the implant but not sure if they are actively secreting enzymes the entire time
146
fibrous encapsulation only pertains to what type of biomaterials
non-degradable
147
what is fibrous encapsulation
it is the maturation of granulation tissue, which is marked by the presence of large blood vessels and collagen
happens around an implant
148
the thickness of fibrous encapsulation depends on __
the degree of injury
location of implant - more motion can lead to thicker encapsulation
shape - edges leads to thicker capsule
degradation - short degradation will lead to no capsule
149
in chronic inflammation __ has to be there for a long time
lymphocytes
150
you see __ in chronic inflammation
granulomas - consists of layers of FBGCs surrounding non-phagocoytosable particles
151
what are the 4 resolutions after implantation
1. encapsulation
2. integration - no fibrous layer, cells from tissue form right next to the implant
3. resorption - if the implant degrades, no capsule will form
4. extrusion - the implant will be forced out
152
repair vs. regeneragtion
repair - the defect is replaced with scar tissue (occurs when cut extends into dermal layer)
regeneration - identical tissue is formed (only occur when injury is in epithelial layer )
153
how to do you tell if something is repair or regeneration in vivo?
you can't. you need to take a sample of the tissue and test it in vitro
154
what are the consideration for in vivo studies for implants
1. choice of animal
2. implant site
3. length of study
4. biomaterial considerations (shape, size)
5. controls
155
what are the types of controls used in studies
contralateral control (opposite uninjured limb) shows level of regeneration
unfilled defect - asses level of improved healing
156
what is responsible for the allergic response
acquired immune response
157
what are the 4 characteristics of acquired immunity
1. specificity - antigen will bind to specific antibody in the humoral response
2. diversity - response to a wide array of thing
3. self/non self recognition - problem with autoimmune disorders
4. memory - this is why vaccines work
158
what are the 2 types of acquired immunity
humoral immunity - action of antibodies
cellular immunity - action of t cells
159
describe humoral immunity
it is the action of antibodies. mediated by B cells
| main way to fight off foreign invaders
160
describe cellular immunity
action of t cells
detect altered self cells
161
the specificity of acquired immunity comes from __
lymphocytes (b and t cells) that are involved in an immune reaction are active in response to antigens
162
what is an antigen
it is a substance that binds to an antibody or t cell receptor to initiate the acquired immune response
antigen has a high molecular weight
163
what is a hapten
it is a low MW substance that combines with a high MW molecule to induce an immune response
upon subsequent exposure, you don't need the larger molecule to induce an immune response
the small molecule can induce an immune response by itself even though it normally wouldn't (metal allergies)
164
what is an adjuvant
it non specifically enhances immune response to antigens
it increases uptake of antigens by phagocytic cells
165
what does the compliment system do
it causes the elimination of foreign elements
it is the bridge between innate immunity and acquired immunity
166
what are the 2 parts of the compliment system
the classical and alternative pathway.
they converge to cause formation of the membrane attack complex to lyse foreign cells
167
how does the classical pathway begin
with the binding of an antibody to an antigen on a bacterial cell
the attachment of the antibody induces a conformational change, which causes a cascade of chemical reactions starting with the cleavage of C4
