bio sensors lecture 1

8/2/2019 Bio Sensors Lecture 1

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BIOSENSORS AND BIOCONJUGATION

BME 140 Bioengineering

Boaz ViloznyOctober 4, 2011


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What is a sensor?

A sensor is any device that converts a measurement into a signal


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What is a biosensor?

A sensor in which the receptor is a biomoleculeA sensor in which the analyte is a biomoleculeA sensor in which the signal is produced bybiological interactions


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Optical Biosensors


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Indicators and immobilized sensors

homepages.wmich.edu/~rossbach/bios312
http://homepages.wmich.edu/~rossbach/bios312/LabProcedures/Phenol%20Red%20Results.htmlhttp://homepages.wmich.edu/~rossbach/bios312/LabProcedures/Phenol%20Red%20Results.html


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Optical biosensor: DNA microarray

What is the receptor?What is the analyte?How is the signal transduced?Is this a true sensor?

http://www.wormbook.org/chapters/www_germlinegenomics


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Fluorescent biosensor: Cameleon

What is the receptor?

What is the reporter?

How is the signal transduced?

How is sensor delivered to cells?

http://probes.invitrogen.com/media/pis/mp36207.pdf


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Electrical Biosensors

What is the electrical signal?PotentiometryAmperometry

VoltammetryConductance, etc.

No optical components

MiniaturizableSensitivity increases with nanofabrication


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Receptors for electrical biosensors

EnzymesAntibodiesDNA/RNA


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Electrical biosensor design

Sandwich Immunoassay

Urban, 2009 Measurement Science and Technology


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Bioconjugation and biosensor fabrication

How is bioconjugation used?Requirements of bioconjugation

SelectiveStablePhysiological conditions

Non-chemical affinity methods

Nucleic acid hybridizationProtein/substrate interactions


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Bioconjugation: chemical crosslinking

Why a chemical bond?Chemistry for special surfaces

Metal oxidesGoldOrganic polymersCharged surfaces


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Optical immunoassay on a chip

Raj, J. et al , Biosens. Bioelectron. 2009

Surface treatment Amino-Silanization Crosslinker Capture antibody Protein-blocking Analyte (IgG) Labeled antibody Optical measurement


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Voltammetric aptamer-biosensor

Zhao, G-C. et al , Electrochem. Commun. 2010

Why not a sandwich assay? How to achieve selectivity? How is signal generated?

Gold surface on glassycarbon electrode

Doxorubicin as indicator:redox active, dsRNAintercalator


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Putting it all together: the biosensoras a self-contained diagnostics lab

http://www.technologyreview.com/biomedicine/38201/

Chin, C.D.; Sia, S.K. et al . Nature Medicine, 2011


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The problem: low-cost diagnostics

Resource-limited settingsLack of medical facilitiesLack of doctors

Lack of moneyHealth epidemicDiagnostics can make a difference

HIV, siphylis treatable; avoids transmission to newborns


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Why bioengineering?

MicrofluidicsBiochemistryNanotechnologyMedicineEngineering


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Antibody detection: ELISA

Immobilized antigen

Target antibody

Gold-labeled secondary antibody


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Signal amplification


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Liquid handling


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Does it work?

70 specimenstested

15 minutes, 1LbloodOne false positive(96% specificity)

Chin, C.D.; Sia, S.K. et al . Nature Medicine, 2011


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Biosensing with nanopipettes
http://nanopipette-fabrication.avi/


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Nanopipettes as sensors

FabricationIonic currentCurrent rectification


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Ion current through nanopipettes


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An electrical DNA nanosensor

Fu, Baker, et al Chem. Commun. 2009

Detected 15-base oligomersSignal detected by current rectification


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A renewable nanosensor

Piper, Klenerman et al , J. Am. Chem. Soc. 2006

Reporter dye trapped inpipette tip

Fluorescence increases inpresence of sodium

Imaged with confocalmicroscope


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Label-free immunosensing withnanopipettes (STING)

Art by David Liao


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Electrical antibody recognition

-20 -10 0 10 20-1.1

-1.0

-0.9

-0.8

0.8

0.9

1.0

1.1

I ,

n o r m a

l i z e

d

Time [min]

+VEGF

Anti-ferritinAnti-VEGF

Umehara, Pourmand et al ,Proc. Nat. Acad. Sci. 2009


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Calcium ions: a good target for biosensing

Major affector of cellularevents

Indicator ofcytotoxycity/apoptosisSingle-cell arrays used tomonitor cellular events bymonitoring cytosolic [Ca2+ ] 1,2

1. Xu, et al. Anal. Biochem. 2010

2. Li et al. Integr. Biol. 2009 Li, 2009


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A receptor for calcium: Calmodulin


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Quartz surfacefunctionalization


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buffer (pH 7)

Ca 2+ (0.1 mM)

buffer

Change bath


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Calcium sensor is reversible

buffer buffer buffer

Ca 2+ Ca 2+


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And selective


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Summary

Nanopipettes modified with Calmodulinas a recepterReversible calcium signalSelective over magnesiumDetection limit 2x10 -5 M Ca2+ .Further work: improve surface chemistry,sensitivity, shelf life.Test with biological systems


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Summary

Biosensors: form and functionBioconjugation: making biomolecules behaveBiosensor case studies

Optical sandwich immunoassayElectrical aptamer sensormCHIP immunoassay

STING bioelectrical sensor

bio sensors lecture 1

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