STUDIES OF DIRECTED ADHESION OF BIO-ORGANISMS ON

126 Pages·2005·1.5 MB·German

STUDIES OF DIRECTED ADHESION OF BIO-ORGANISMS ON
FUNCTIONALIZED POLYDIMETHYLSILOXANE ELASTOMER

By

THOMAS G. ESTES

A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY

UNIVERSITY OF FLORIDA

2005
Copyright 2005

by

Thomas G. Estes
This document is dedicated to my future wife Nancy Goodman for all that she means to
me and for all of her motivation and support.
ACKNOWLEDGMENTS
The Lord has blessed me with supportive parents, siblings, support group and a
future wife that have helped me accomplish my goals. For that I am forever grateful. I
would like to thank my parents for telling me that I could accomplish any goal I set. The
relationships I acquired at The Citadel taught me so many wonderful things and have
given me an extended family that is irreplaceable. The community of Gainesville with
the families of Westside Baptist Church has been a great support group for me while
attending graduate school. Thank you is in order for Kevin R. Griffin for teaching me
what it means to be a man.
I am also Grateful to Dr. Brennan for believing in me and the other faculty and staff
members that have helped me: Dr. Ambrose, Dr. Baney, Dr. Batich, Dr. Goldberg, Dr.
Holloway, Dr. Mecholsky, Dr. Segal, Dr. Holm, Dr. Rittschof, Dr. Maurene Callow, Dr.
Jim Callow, Dr. Wendt, Dr. Finlay, Elaine Beem, Denise Watson, Greg Kowalke, Beatriz
Orihueala, Laura Sautina and Jennifer Wrighton. The Brennan, Batich and Goldberg
Research Groups have helped me each step of the way. I would like to thank: Dr. Leslie
Wilson, Dr. Cliff Wilson, Dr. Clay Bohn, Dr. Charles Seegert, Dr. Adam Feinberg, Dr
Brian Hatcher, Dr. Nikhil Kotukar, James Schumacher, Michelle Carman, Amy Gibson,
Kiran Karve, Kennith Williams, Matthew Blackburn, Chris Long, Dr. Joshua Stopek, Dr.
Chris Widenhouse, Dr. Brett Almond, Dr. Brian Cuevas, Dr. Adam Reboul, Dr. Lynn
Peck, Dr. Dan Urbaniak, D.r Margaret Kayo, Amanda York, Paul Martin, Iris
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Schumacher, Shema Freeman, Amin Elachchabi, Samesha Barnes, Dr. Brad Willenberg,
Taili Thula, Jompo Moleye.
I would also like to thank the staff at the Major Analytical Instrumentation Center
(MAIC) for heir assistance, Dr. Dempere, Wayne Acre and Eric. Thank you is also in
order to the Office of Naval Research for funding me. I also would like to thank Dr.
Cherie Everhart at Kimberly-Clark for giving me my first job and her husband Dr. Denis
Everhart for his assistance.
I would also like to thank the Citadel men who gave their lives serving our country
over the past few years: Richard Childress, Dan Malcolm and Drew Mazur.

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TABLE OF CONTENTS

page
ACKNOWLEDGMENTS.................................................................................................iv
LIST OF TABLES...............................................................................................................x
LIST OF FIGURES...........................................................................................................xi
ABSTRACT.......................................................................................................................xv
CHAPTER
1 INTRODUCTION........................................................................................................1
Biological Adhesion.....................................................................................................2
Marine Biofouling........................................................................................................2
Cardiovascular Disease.................................................................................................3
Proposed Research........................................................................................................4
Experimental Summary................................................................................................5
2 BACKGROUND..........................................................................................................7
Introduction...................................................................................................................7
Wettability and Adhesion.............................................................................................7
Wettability.............................................................................................................7
Adhesion................................................................................................................9
Relating Wettability and Adhesion.....................................................................10
Biological Adhesion...................................................................................................10
Topographical Studies.........................................................................................11
Protein and Antibody Grafting............................................................................13
Polydimethylsiloxane Elastomer................................................................................14
Surface Modulus Modifications..........................................................................16
Surface Chemical Modifications.........................................................................16
Plasma Treatment.........................................................................................17
Surface Topgraphical Modification.....................................................................19
Cure Conditions..........................................................................................................19
Current Uses of PDMSe.............................................................................................20
Marine Biofouling......................................................................................................20
Microfoulers........................................................................................................20
Macrofoulers........................................................................................................21
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Marine Biofouling Applications.................................................................................23
Biomedical Biofouling and Cardiovascular Disease..................................................24
Biomedical Applications............................................................................................25
Microfluidic Devices...........................................................................................25
Circulating Endothelial Cells..............................................................................26
Endothelial Progenitor Cells................................................................................26
Proposed Research......................................................................................................27
3 FIELD ASSAYS INVESTIGATING BARNALCE AND BRYOZOAN
SETTLEMENT ON SURFACE MODIFIED POLYDIMETHYLSILOXANE
ELASTOMER............................................................................................................29
Introduction.................................................................................................................29
Experimental Summary.......................................................................................30
Experimental...............................................................................................................30
PDMSe................................................................................................................30
Allyltriethoxysilane Treatment............................................................................31
Surface Modification...........................................................................................31
Film thickness..............................................................................................31
Plasma treatment..........................................................................................32
Topographical Modification................................................................................33
Characterization...................................................................................................34
Field Setup...........................................................................................................35
Results and Conclusions.............................................................................................35
Characterization...................................................................................................35
Hydrophobic recovery..................................................................................35
Contact angle................................................................................................36
Field Study 6-1-03: Settlement of Balanus amphitrite and Bugula neritina.......37
Barnacle settlement......................................................................................37
Bryozoan settlement.....................................................................................37
Field Study 7-3-03: Settlement of Balanus amphitrite and Bugula neritina.......38
Barnacle settlement......................................................................................38
Bryozoan settlement.....................................................................................39
Conclusion..................................................................................................................40
4 LAB ASSAYS INVESTIGATING THE EFFECTS OF PLASMA TREATMENT
AND CURE CONDITIONS OF POLYDIMETHYLSILOXANE ELASTOMER
ON BARNACLE AND ULVA SPORE SETTLEMENT AND RELEASE..............42
Introduction.................................................................................................................42
Experimental...............................................................................................................44
Bioassays.............................................................................................................46
Barnacle assays............................................................................................46
Spore assays.................................................................................................48
Results and Discussion...............................................................................................48
Barnacle Assays...................................................................................................48
Spore Assays.......................................................................................................51
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Conclusions.................................................................................................................52
5 LAB ASSAYS INVESTIGATING THE EFFECTS OF TOPOGRAPHICAL
MODIFICATION OF POLYDIMETHYLSILOXANE ELASTOMER ON
BARNACLE SETTLEMENT AND RELEASE........................................................54
Introduction.................................................................................................................54
Experimental...............................................................................................................57
Sample Preparation..............................................................................................57
Lab Experiment...................................................................................................58
Results and Discussion...............................................................................................59
Conclusion..................................................................................................................62
6 IN VITRO ASSAYS INVESTIGATING SPECIFIC BINDING OF PROTEINS
ANTIBODIES AND ENDOTHELIAL CELLS TO SURFACE MODIFIED
POLYDIMETHYLSILOXANE ELASTOMER........................................................63
Introduction.................................................................................................................63
Experimental...............................................................................................................67
Experimental Overview.......................................................................................68
Experimental Details...........................................................................................69
PMDSe film production...............................................................................69
KOH and HCl treatments.............................................................................69
Aminopropyltriethoxy silane treatment.......................................................70
Protein Addition to PDMSe.........................................................................70
Absorbance assays........................................................................................73
Fluorescence assays......................................................................................74
Statistics.......................................................................................................75
Results and Discussion...............................................................................................75
Conclusions.................................................................................................................89
7 FUTURE STUDIES...................................................................................................91
Introduction.................................................................................................................91
Marine Biofouling......................................................................................................91
Biomedical Adhesion..................................................................................................93
APPENDIX
A PROTOCOL FOR RADIO FREQUENCY GLOW DISCHARGE (RFGD)
ARGON (Ar) PLASMA TREATMENT OF POLYDIMETHYLSILOXANE
ELASTOMER CREATED BY: THOMAS ESTES, (REVISED 10/15/03)..............96
Materials Required......................................................................................................96
Equipment Required...................................................................................................96
Methods......................................................................................................................96
Step 1 Turning on all the Parts of this Setup.......................................................96
Step 2 Bell Jar Cleaning and Practice Run..........................................................97
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Step 3 Plasma Treatment.....................................................................................98
Step 4 Turning off the System.............................................................................98
LIST OF REFERENCES...................................................................................................99
BIOGRAPHICAL SKETCH...........................................................................................109

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LIST OF TABLES
Table page

4-1. Elastic modulus and roughness values of PDMSe.....................................................43
4-2. Data table for breakage of barnacles versus treatment...............................................49
6-1. Concentrations of BSA in PBS for standard curve.....................................................72
6-2. Concentrations of mouse IgG in PBS for binding......................................................72
6-3. Concentrations of BSA in PBS for a 10 µg/ml blocking solution..............................72
6-3. BCA reagent components...........................................................................................73

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