1. SERS Detection in Complex Matrices Haynes group in collaboration with Lili He, Srinand Sreevatsan, and Ted Labuza (Department of Food Science)

2. Why is SERS detection in complex 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)

3. Antibody-based SERS detection SERS Detection in Complex Matrices Allowing analysis within co-cultures that simulate in vivo conditions. Ag dendrites 1. Adsorb protein G 2. Attach anti-OVA 3. Capture OVA LaserSERS SERS shift L. He, et al., Analytical Chemistry, 2011 http://chemistry.umeche.maine.edu/CHY431/Conformation5.html Ovalbumin: egg white protein Potentially allergenicPotentially allergenic Proof-of-concept protein for generalizableProof-of-concept protein for generalizable assay development assay development immunoglobulin-binding protein from Streptococcal bacteria

4. Ag Dendrite SERS Substrate Ag dendrite UV-Vis spectrum Zn + 2AgNO 3 2Ag + Zn(NO 3 ) 2 L. He, et al. Journal of Raman Spectroscopy, 2009.

5. OVA Detection in Milk Raman Shift (cm -1 ) 18001600140012001000800 17001600150014001300 Raman Intensity (AU) 2 nd Derivative Transformation L. He, et al., Analytical Chemistry, 2011 within class standard deviation

6. PCA of Ovalbumin Spectral Data L. He, et al., Analytical Chemistry, 2011 PC1 Score x 10 PC2 Score x 10 -0.20.3 -0.3 0.2 Ag dendrites Ag-protein G Ag-G-anti-OVA Ag-G-anti-OVA-OVA

7. OVA Detection in Milk L. He, et al., Analytical Chemistry, 2011 PBSMilk PC1 Score PC2 Score PC1 Score PC2 Score 0 µg/mL OVA 0.1, 0.5, and 1 µg/mL OVA 0 and 1 µg/mL OVA 5 µg/mL OVA

8. DNA Aptamers as Capture Reagents http://aptamer.icmb.utexas.edu/index/php  Oligonucleotide sequences that bind to a target  Generated by repeated selection cycles  Relatively easy to manipulate  Raman spectra of DNA differ significantly from protein spectra

9. 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/ricin E. Rutenber, et al. Proteins: Structure, Function, and Genetics, 1991 - Aptamer-Based SERS Assay L. He, et al., Chemical Science, 2011

10. - Aptamer-Based SERS Assay L. He, et al., Chemical Science, 2011 Raman shift (cm -1 ) Assignments Aptamer (*) 1316 Adenine (ring stretching) 1022 DNA backbone(PO 2 - stretching) 792thymine, cytosine 735Adenine (ring breathing) Ricin B chain 985 Phenylalanine (ring breathing) 716 Cysteine (C-S stretching) 621 Cysteine (C-S stretching) Raman Intensity (AU) Ag-Ap Ag-Ap-Ricin Ag-Ricin 985 cm -1 621 cm -1 Ag-Ap Ag-Ap-Ricin Ag-Ricin Ag-Ap Ag-Ap-Ricin Ag-Ricin * * * *

11. - Aptamer-Based SERS Assay L. He, et al., Chemical Science, 2011 PC1 Score PC2 Score -1619 -811 Future work: capture analyte closer to SERS-active surface to improve signal

12. M Aptamer Capture in Food Matrices L. He, et al., Chemical Science, 2011 Compare to 5  g/mL using antibody approach