Holographic Angular Domain Elastic Scattering of Single Biological Cells

227 Pages·2014·7.14 MB·English

Holographic Angular Domain Elastic
Scattering of Single Biological Cells
by
Dustin W. Shipp
Submitted in Partial Fulfillment
of the
Requirements for the Degree
Doctor of Philosophy
Supervised by
Professor Andrew J. Berger
The Institute of Optics
Arts, Sciences and Engineering
Edmund A. Hajim School of Engineering and Applied Sciences
University of Rochester
Rochester, New York
2014

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Tomythreegirls: Laura,ElianaandChristine.
YouarethemostwonderfulsourcesoflightIcouldimagine.
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Biographical Sketch
Dustin W. Shipp was born in Oklahoma City, Oklahoma, but spent most of his
childhoodinSt. Louis,Missouri. HeattendedBrighamYoungUniversityinProvo,
UT, where he was awarded the Gordon B. Hinckley Presidential Scholarship. His
studieswereputonholdfortwoyearsasheservedasamissionaryfortheChurchof
JesusChristofLatter-daySaintsinFrankfurt,Germany. In2008,hegraduatedwith
BachelorofSciencedegreesinPhysicsandMathematics. Hethenbeganhisdoctoral
studies in Optics at the University of Rochester that same year. As a doctoral
student, he pursued research in biomedical spectroscopy - including Raman and
angular scattering - under the direction of Dr. Andrew J. Berger. The Center for
ExcellenceinTeachingandLearning(CETL)attheUniversityofRochesterawarded
himacertificateinAdvancedTeaching.
BIOGRAPHICALSKETCH iv
Thefollowingpublicationsarearesultofworkconductedduringdoctoralstudy:
D. W. Shipp, R. Qian, A. J. Berger, “Angular-domain scattering interferometry,”
OpticsLetters 38(22),pp. 4750–4753,2013.
ff
D. W. Shipp, S. Mitra, T. H. Foster, and A. J. Berger, “E ect of photodynamic
therapyonsinglecancercellsstudiedbyintegratedRamanandangularscattering
microscopy,”ProceedingsofSPIE8219,82190G,2012.
D.W.ShippandA.J.Berger,“Time-lapsedintegratedRamanandangularscatter-
ingmicroscopyofimmunecells,”ProceedingsofSPIE7907,79070X,2011.
Thefollowingpresentationsweregivenbasedonworkconductedduringdoctoral
study:
OralPresentation-D.W.Shipp,R.Qian,AshleyE.Cannaday,andA.J.Berger,“Pre-
cise, time-lapsed measurements of organelle sizes in single cells by Holographic
AngularDomainElasticScattering,” SPIEPhotonicsWest,February2,2014.
Poster Presentation - D. W. Shipp, R. Qian, and A. J. Berger, “Angular-domain
scattering interferometry,” ECI Advances in Optics for Biotechnology, Medicine, and
Surgery,June2-5,2013.
Oral Presentation - D. W. Shipp, R. Qian, and A. J. Berger, “Size- and position-
dependent angular scattering interferometry,” SPIE Photonics West, February 3,
2013.
ff
Oral Presentation - D. W. Shipp, S. Mitra, T. H. Foster, and A. J. Berger, “E ect
of photodynamic therapy on single cancer cells studied by integrated Raman and
angularscatteringmicroscopy,”SPIEPhotonicsWest,January22,2012.
Poster Presentation - D. W. Shipp, D. C. Davidson, M. Kiebala, S. B. Maggirwar,
and A. J. Berger, “Platelet activation studied by Raman and angular scattering mi-
croscopy,”WorldAIDSDayatURMC,December1,2011.
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Oral Presentation - D. W. Shipp and A. J. Berger, “Time-lapsed integrated Raman
andangularscatteringmicroscopyofimmunecells,”SPIEPhotonicsWest,January
23,2011.
Oral Presentation - D. W. Shipp and A. J. Berger, “Time-lapsed integrated Raman
and angular scattering microscopy of immune cells,” OSA Frontiers in Optics, Oc-
tober27,2010.
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Acknowledgments
Several individuals have played crucial roles in the completion of this thesis. I
wouldliketoacknowledgeandexpressmygratitudetoeachofthem.
I am grateful for the support and guidance of my thesis advisor, Prof. Andrew
Berger. Hislong-termvisionandcarefulattentiontodetailenabledustoovercome
the series of hurdles we encountered and create a unique and useful tool. I am
especially grateful for his emphasis on advising, training, and educating that has
provided me with a foundation on which I can build the rest of my career. This
emphasisalsoservesasanexampleinmymentoringandteaching.
I had the privilege of having my thesis research supported by a brilliant and
generous thesis committee consisting of Profs. Miguel Alonso, Tom Brown, and
Tom Foster. Each of them was very willing to lend their time and respective ex-
pertise to the project. Prof. Alonso’s knowledge of coherence was instrumental
in understanding the speckle in our system and various ways to remove it. Prof.
Brown was very helpful as we developed our interferometer and began to inter-
pret the measurements it acquired. Prof. Foster was incredibly generous with his
time and laboratory supplies to provide cells for our biological experiments. He
ACKNOWLEDGEMENTS vii
and other members of the his group were very helpful over years of cell culture
troubleshooting,especiallyDrs.TimBaranandSoumyaMitra.
In Prof. Berger’s research group, I was fortunate to be surrounded by many
gifted scientists. Dr. Zachary Smith provided my first introduction to the group
and entrusted me with the IRAM instrument of his creation. Even years after
he graduated from the group, he was happy to help solve some of the system’s
“quirks.” Drs. Brooke Beier and Jason Maher served as my guides entering the
cultureofbiomedicalopticsresearch. Therestofmycareerwillbeshapedbytheir
advice,examples,andfriendship.
The final stages of this work were carried out alongside a fantastic group of
researchers in the Berger group. Much of this research would not have happened
without the help of Ashley Cannaday culturing and measuring cells, Ruobing
(Frank) Qian measuring beads, platelets, and countless other samples, Janet Sor-
rells and Nick Giglio perparing fluorescence measurements, and Addie Stavlo
honingthedataprocessingalgorithms. Iwasfortunatetobeabletodrawontheir
numeroustalentsandandtocallthempartofourgroupfamily.
IamalsogratefulforthesupportfrommanythroughouttheInstituteofOptics.
Whenever I encountered questions beyond my area of expertise, I was easily able
tofindfriendswhoareexpertsinthatfield. IwouldespeciallyliketothankAlden
Jurling and Dustin Moore from the lab of Prof. Jim Fienup for helpful discussions
on phase and speckle. Per Adamson was always willing to give up a piece of
ACKNOWLEDGEMENTS viii
equipment or a couple hours of his time to help solve some mystery in the lab.
The continued existence of our research was made possible by the support of the
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Institute’s administrative sta including Betsy Benedict, Kari Brick, Lissa Cotter,
GinaKern,LoriRussel,GayleThompson,andNoeleneVotens. Iamalsofortunate
tohavefoundmanyfriendshipsinandoutoftheInstituteduringthistime.
Finally, I want to acknowledge the love, support, motivation, and joy I have
receivedfrommyfamily. Iowemanysmilestomydaughters,ElianaandChristine.
I owe many more to my best friend and wife, Laura. Thank you for inspiring me,
calmingme,andforfillingmylifewithhappiness.
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Abstract
The size of organelles can indicate many things about the health or behavior
of cells. Elastic light scattering provides a promising method to non-invasively
measure organelle sizes in living cells. Angle-resolved light scattering of multi-
cell suspensions or tissues has been used to diagnose dangerous pre-cancerous
conditionsandtrackchangesinsub-cellularstructureduringapoptosis.
Angularscatteringmeasurementsofsinglecellswouldallowresearcherstoex-
ff
aminedi erencesinstructureandbehavioramongindividualcells. Unfortunately,
ffi
singlecellscatteringmeasurementsaremoredi cultbecausethesmallilluminated
region leads to larger speckle grains. This speckle causes organelle size estimates
tobeinaccurateandunstable.
This thesis presents a holographic angular domain elastic scattering (HADES)
systemthatinterferometricallymeasuresthefullcomplexfieldofscatteredlight. It
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alsodescribesmultiplemethodsofusingthiscomplexfieldtoovercomethee ects
ofspeckleandobtainscatterer-sizeestimates. Twomethodsarepresentedthatare
appropriateforanalyzingensemblesofdiscretescatterers(e.g.,micro-beads). Sub-
cellularstructureismorecontinuous,soanothermethodisusedtoreducespeckle