Table Of ContentMethods in
Molecular Biology 2394
LLuucciiffeerraassee
Avraham Rasooly AAPPCC
Houston Baker TTooxxiinn
TT--CCeellll
Miguel R. Ossandon Editors
Biomedical
Engineering
Technologies
Volume 2
M M B
ETHODS IN OLECULAR IO LO GY
SeriesEditor
JohnM.Walker
School of Lifeand MedicalSciences
University ofHertfordshire
Hatfield, Hertfordshire, UK
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For over 35 years, biological scientists have come to rely on the research protocols and
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Biomedical Engineering
Technologies
Volume 2
Edited by
Avraham Rasooly
Division of Cancer Treatment and Diagnosis, National Cancer Institute, Rockville, MD, USA
Houston Baker
Division of Cancer Treatment and Diagnosis, National Cancer Institute, Rockville, MD, USA
Miguel R. Ossandon
Division of Cancer Treatment and Diagnosis, National Cancer Institute, NIH, Bethesda, MD, USA
Editors
AvrahamRasooly HoustonBaker
DivisionofCancerTreatment DivisionofCancerTreatment
andDiagnosis andDiagnosis
NationalCancerInstitute NationalCancerInstitute
Rockville,MD,USA Rockville,MD,USA
MiguelR.Ossandon
DivisionofCancerTreatment
andDiagnosis
NationalCancerInstitute,NIH
Bethesda,MD,USA
ISSN1064-3745 ISSN1940-6029 (electronic)
MethodsinMolecularBiology
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https://doi.org/10.1007/978-1-0716-1811-0
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v
Preface
ThisistheThirdEditionofBiosensorsandBiodetection:MethodsandProtocolspublishedin
the Springer Protocols Methods in Molecular Biology series. Biosensing, bioengineering,
and biomedical technologies are discussed from an experimental point of view alongside
detailed protocols useful in developing similar devices. While the first (volumes 503–504)
and second (volumes 1571–1572) editions of the Methods in Molecular Biology series
focused on biosensor technologies, in this edition we expanded the scope of subjects to
includenon-sensorbiomedicaltechnologies.
This edition complements and broadens the previous editions by presenting new
technologies not included in previous editions which are still relevant today. For example,
the Kretschmann configuration for surface plasmon resonance (SPR) or the fabrication of
variouselectrochemicalelectrodesiswelldescribedinthepreviouseditions,andwereferthe
reader tothepreviouseditions.Thethirdeditionincludestwovolumes:
Volume 1 Biosensors and Imaging Technologies provides experimental protocols on
biosensorsandmedicalimagingtechnologiessuchastomography,MRI,andNMR.
Volume 2 Biomedical Technologies is focused on biomedical technologies including
molecular and cellular analytical methods, experimental new drug delivery approaches,
guidedsurgery,implants,andtissueengineering.
A biosensor is defined by the International Union of Pure and Applied Chemistry
(IUPAC) as “A device that uses specific biochemical reactions mediated by isolated enzymes,
immunosystems, tissues, organelles or whole cells to detect chemical compounds usually by
electrical, thermal or optical signals” [1]. All biosensors are based on a two-component
system:
1. Biological recognition element(s) (ligands) such as an enzyme, antibody, DNA,or cell
that facilitates a specific binding or biochemical reaction with the target analyte. The
liganddeterminesthespecificityofthebiosensor.
2. Signal conversion unit (transducer) which converts ligand binding into a measurable
signal(e.g.,electrical,optical,piezoelectric).
Dr.LelandCClarkpioneeredresearchonbiosensorsintheearly1960sanddeveloped
the“enzymeelectrode”conceptforglucoseconcentrationmeasurementusingtheenzyme
Glucose Oxidase (GOD) [2], thereby incorporating both ligand and transducer. Glucose
monitoringisessentialfordiabetespatients.Eventodaythemostcommonclinicalbiosensor
technologyforglucoseanalysisistheelectrochemicaldetectionmethodenvisionedbyClark
morethan60yearsago.Currentglucosemonitoringisperformedusingrapidpointofcare
biosensors made possible through advances in electronics that have enabled sensor minia-
turizationandsimple,rapid,andaccurateanalysis.
Biosensing Modalities
1. Direct (label-free) recognition sensors in which the biological interaction is directly
measured by the transducer (e.g., change of refraction index) without any enzymatic
orbiochemicalassay.
vii
viii Preface
2. Indirectdetectionsensorswhichrelyonlabelingutilizingsecondaryelements.Themost
commonlyusedsecondaryelementsareenzymes(usuallycatalytic)andfluorescenttags.
Ligandsaremoleculesthatbindspecificallywiththetargetmoleculetobedetected.The
most important properties of ligands are affinity and specificity. Of the various types of
ligandsusedinbiosensors,immunosensors—particularlyantibodies—arethemostcommon
biosensor recognition element. Antibodies (Abs) are highly specific and versatile and bind
stronglyandstablytospecificantigens.However,Abligandshavelimitedlong-termstability
andaredifficulttoproduceinlargequantitiesformulti-targetbiosensorapplicationswhere
manyligandsareneeded.
Other types of ligands such as aptamers and peptides are more suited to high-
throughput screening and chemical synthesis. Aptamers are protein-binding nucleic acids
(DNAorRNA)selectedfromrandompoolsbasedontheirabilitytobindothermolecules
with high affinity. Peptides are another potentially important class of ligand suitable for
high-throughputscreeningduetotheireaseofselection.However,theaffinityofpeptidesis
often lower than that of antibodies or aptamers, and peptides vary widely in structural
stabilityandthermalsensitivity.
Volume 2 includes two parts: (1) Molecular and Cellular Technologies and (2) Thera-
peutics Technologies. This volume focuses on experimental technologies developed to
address medical and health-related topics. Several technologies are applicable to the study
of cancer etiology, progression, metastasis and recurrence, as well as the development of
noveltherapeuticapproachesandassessmentoftheefficacyofcancer therapies.
Molecular and Cellular Technologies: The chapters in this part fall into three tech-
nology research areas: (1) cellular analysis, (2) molecular analysis, and (3) molecular and
cellular manipulation.
1. Cellular analysis technologies includes chapters focusing on tumor cells and tumor-
igenesis, with topics such as analysis of circulating tumor cells, dormant cancer cells,
microtumors,andthetumormicroenvironment.Otherchaptersoncelllineagetracing
or imaging probes for cell-based measurements are also relevant for analysis of disease
progression and therapeuticresistance.A major themewithinthissubsection issingle-
cellanalysis,whichhassignificantclinicalpotentialforearlydetectionandmayprovide
information on the nature and heterogeneity of a disease, disease progression, or
resistance to treatment. Several chapters describe single-cell analysis technologies such
as microfluidic devices, single-cell subtype analysis, or real-time analysis of signaling at
thesingle-celllevel.
2. Molecular analysis technologies includes several different molecular technologies,
including droplet technologies, which can be applied to DNA manipulation and for
direct delivery of nucleic acid therapeutics into cells. Also included are various single-
molecule localization microscopy approaches for detecting receptors and assessing the
binding capacity for molecular targets. When combined with analytical technologies
suchas microchip free-flow electrophoresis, green chemistryextraction for multi-omic
analyses, or direct in situ labeling of DNA double-stranded breaks, these approaches
provide powerful tools for elucidating molecular mechanisms and understanding dis-
easeprogression.
3. Molecular and cellular manipulation technologies incorporates a broad array of
technologies, such as those to incorporate noncanonical amino acids in yeast, use of
opticaltweezerstodissectallostericcommunicationnetworks,nuclease-assistedminor-
Preface ix
allele enrichment, and technologies for lossless ion manipulations. The innovative
technologies described in these chapters offer new approaches to the study of cancer
cellbiology.
The second part of Volume 2 describes therapeutic technologies and is divided into
three areas: (1) drug delivery (2) surgical and therapeutics devices, and (3) implants and
tissueengineeringtechnologies.
1. Drugdeliverytechnologiesprovidesprotocolsforseveraldifferentapproachesinclud-
ing ultrasound-mediated drug delivery, fluorescent mapping readout of drug delivery,
and nanostructure-based drug delivery using nanoparticles, microbubbles, and
nanovesicles.
2. Surgery and therapeutic devices describes a wide range of therapeutic technologies
ranging from cancer stem cell targeting with bacteriophage and fluorescence-guided
tumorresectiontoX-ray-inducedphotodynamictherapyandthermalablation.Several
chapters cover tumor targeting technologies such as fluorescence monitoring of drug-
target engagement, fluorescence in vivo imaging of tumors, and a porcine model to
evaluateoncologicaltherapies.
3. Implantsandtissueengineeringtechnologiesisthefinalsubsectionofthebookand
includes protocols for utilizing natural material such as avian lungs for pulmonary
therapeutics,collagenforhumanlymphaticmicrovessels,ornaturalpolymer(hydroxy-
apatite, collagen, cellulose acetate) scaffolds for bone tissue engineering. This subsec-
tionalsoincludestechnologiestomonitorimplantssuchasmeasurementofneutrophil
accumulationonbiomaterial,imagingofimplantedhyaluronicacidhydrogelbiodegra-
dation, bio-tribometer assessment of orthopedic metal implant debris, and computa-
tionalmodelingofobstructionsontheperformanceofventricularcatheters.
Thisbookdescribesthebasictypes,designs,andapplicationsofbiosensorsandimaging
technologiesfromanexperimentalpointofview.Wehaveassembledmanuscriptsrepresent-
ing themajor technologies in the field and haveincluded enough technical information so
that the reader can both understand the technology and carry out the experiments
described.
The target audience for this book includes medical, engineering, and biomedical
researcherswhoaredevelopingbiomedicaltechnologies.Othertargetgroupsarebiologists
andclinicianswhoultimatelybenefitfromdevelopmentandapplicationofthetechnologies.
In addition to research scientists, the book may also be useful as a teaching tool for
bioengineering,biomedicalengineering,andbiologyfacultyandstudents.Tobetterrepre-
sent the field, most topics are described in more than one chapter. The purpose of this
redundancyistobringseveralexperimentalapproachestoeachtopic,toenablethereaderto
chooseanappropriatedesign,tocombineelementsfromdifferentdesignsinordertobetter
standardizemethodologies,andtoprovidereadersmoredetailedprotocols.
Rockville,MD,USA MiguelR.Ossandon
HoustonBaker
AvrahamRasooly
x Preface
References
1. IUPAC Compendium of Chemical Terminology 2nd Edition (1997). (1992), Interna-
tionalUnionofPureandAppliedChemistry:ResearchTrianglePark,NC,U.S.
2. Clark LC Jr, Lyons C (1962) Electrode systems for continuous monitoring in cardio-
vascularsurgery.AnnNYAcadSci102:29–45.kl
