SERS Detection in Complex MatricesSERS Detection in Complex Matrices

Haynes group in collaboration with Lili He, Srinand Sreevatsan, and Ted Labuza

(Department of Food Science)

Why is SERS detection in complex Why is SERS detection in complex matrices important?matrices important?

• Intentional or accidental adulteration of food supply– Equipment contamination by allergens– Bioterrorism at vulnerable points of food supply chain

• Need rapid, simple method of detecting foreign proteins in food supply– Aromatic amino acid bands ~950 and 1400 cm-1 shift– Amide I and III bands at 1600-1700 and 1200-1350 cm-1

shift, respectively

• Usually done via ELISA or chromatography– Expensive– Slow– Done in lab (rather than on-site)

Antibody-based SERS detectionAntibody-based SERS detection

SERS Detection in Complex Matrices SERS Detection in Complex Matrices

Allowing analysis within co-cultures that simulate Allowing analysis within co-cultures that simulate in vivo in vivo conditions. conditions.

Ag dendritesAg dendrites

1. Adsorb protein G1. Adsorb protein G 2. Attach anti-OVA2. Attach anti-OVA 3. Capture OVA3. Capture OVA

LaserLaser SERSSERS SERS shiftSERS shift

L. He, et al., L. He, et al., Analytical ChemistryAnalytical Chemistry, 2011, 2011http://chemistry.umeche.maine.edu/CHY431/Conformation5.htmlhttp://chemistry.umeche.maine.edu/CHY431/Conformation5.html

Ovalbumin: egg white proteinOvalbumin: egg white protein

•Potentially allergenicPotentially allergenic•Proof-of-concept protein for generalizableProof-of-concept protein for generalizable assay developmentassay development

immunoglobulin-binding protein from immunoglobulin-binding protein from StreptococcalStreptococcal bacteria bacteria

Ag Dendrite SERS SubstrateAg Dendrite SERS Substrate

Ag dendrite UV-Vis Ag dendrite UV-Vis spectrumspectrum

Zn + 2AgNOZn + 2AgNO33 2Ag + Zn(NO 2Ag + Zn(NO33))22

L. He, et al. L. He, et al. Journal of Raman Spectroscopy, Journal of Raman Spectroscopy, 2009.2009.

OVA Detection in MilkOVA Detection in Milk

Raman Shift (cmRaman Shift (cm-1-1))18001800 16001600 14001400 12001200 10001000 800800

Raman Shift (cmRaman Shift (cm-1-1))17001700 16001600 15001500 14001400 13001300

Ram

an

Inte

nsity

(A

U)

Ram

an

Inte

nsity

(A

U)

22ndnd Derivative Derivative TransformationTransformation

L. He, et al., L. He, et al., Analytical ChemistryAnalytical Chemistry, 2011, 2011

within class standard deviationwithin class standard deviation

PCA of Ovalbumin Spectral DataPCA of Ovalbumin Spectral Data

L. He, et al., L. He, et al., Analytical ChemistryAnalytical Chemistry, 2011, 2011

PC1 Score x 10PC1 Score x 10

PC

2 S

core

x 1

0P

C2

Sco

re x

10

-0.2-0.2 0.30.3

-0.3

-0.3

0.2

0.2

Ag dendritesAg dendrites

Ag-protein GAg-protein G

Ag-G-anti-OVAAg-G-anti-OVA

Ag-G-anti-OVA-OVAAg-G-anti-OVA-OVA

OVA Detection in MilkOVA Detection in Milk

L. He, et al., L. He, et al., Analytical ChemistryAnalytical Chemistry, 2011, 2011

PBSPBS MilkMilk

PC1 ScorePC1 Score

PC

2 S

core

PC

2 S

core

PC1 ScorePC1 Score

PC

2 S

core

PC

2 S

core

0 µg/mL OVA0 µg/mL OVA

0.1, 0.5, and 1 µg/mL OVA0.1, 0.5, and 1 µg/mL OVA

0 and 1 µg/mL OVA0 and 1 µg/mL OVA

5 µg/mL OVA5 µg/mL OVA

DNA Aptamers as Capture ReagentsDNA Aptamers as Capture Reagents

http://aptamer.icmb.utexas.edu/index/phphttp://aptamer.icmb.utexas.edu/index/php

Oligonucleotide sequences that bind to a targetOligonucleotide sequences that bind to a target Generated by repeated selection cyclesGenerated by repeated selection cycles Relatively easy to manipulateRelatively easy to manipulate Raman spectra of DNA differ significantly from protein spectraRaman spectra of DNA differ significantly from protein spectra

Target = ricin (potential bioterrorism agent)Target = ricin (potential bioterrorism agent)•No antidoteNo antidote•Need rapid assay for detection in food suppliesNeed rapid assay for detection in food supplies•A-chain and B-chain (non-toxic B chain used to develop assay)A-chain and B-chain (non-toxic B chain used to develop assay)

www.bt.cdc.gov/agent/ricinwww.bt.cdc.gov/agent/ricinE. Rutenber, et al. Proteins: Structure, Function, and Genetics, 1991E. Rutenber, et al. Proteins: Structure, Function, and Genetics, 1991

Aptamer--Based SERS Assay

L. He, et al., L. He, et al., Chemical Science,Chemical Science, 2011 2011

Aptamer--Based SERS Assay

L. He, et al., L. He, et al., Chemical Science,Chemical Science, 2011 2011

  Raman shift (cm-1) Assignments

Aptamer (*)

1316 Adenine (ring stretching)

1022DNA backbone(PO2

- stretching)

792 thymine, cytosine

735 Adenine (ring breathing)

Ricin B chain

985 Phenylalanine (ring breathing)

716 Cysteine (C-S stretching)

621 Cysteine (C-S stretching)

Ram

an In

tens

ity (

AU

)R

aman

Inte

nsity

(A

U)

Ram

an In

tens

ity (

AU

)R

aman

Inte

nsity

(A

U)

Ag-ApAg-Ap

Ag-Ap-RicinAg-Ap-Ricin

Ag-RicinAg-Ricin 985 cm985 cm-1-1621 cm621 cm-1-1

Ag-ApAg-Ap

Ag-Ap-RicinAg-Ap-Ricin

Ag-RicinAg-Ricin

Ag-ApAg-Ap

Ag-Ap-RicinAg-Ap-RicinAg-RicinAg-Ricin

**

******

Aptamer--Based SERS Assay

L. He, et al., L. He, et al., Chemical Science,Chemical Science, 2011 2011

PC1 ScorePC1 Score

PC

2 S

core

PC

2 S

core

-16-16 1919

-8-8

1111

Future work: capture analyte Future work: capture analyte closer to SERS-active surface to closer to SERS-active surface to improve signalimprove signal

Aptamer Capture in Food MMatrices

L. He, et al., L. He, et al., Chemical Science,Chemical Science, 2011 2011

Compare to 5 Compare to 5 g/mL g/mL using antibody approachusing antibody approach