Tip-enhanced Raman Tip-enhanced Raman spectroscopy of single RNA spectroscopy of single RNA

strandsstrands

Elena Bailo and Volker DeckertElena Bailo and Volker DeckertISAS – Institute for Analytical SciencesISAS – Institute for Analytical Sciences

Dortmund, GermanyDortmund, Germany

Poltersdorf, 03-05.10.2007

MotivationMotivation

Direct and label-free sequencing of Direct and label-free sequencing of single DNA strandsingle DNA strand

Characterization of the structure and Characterization of the structure and dynamic of cell wallsdynamic of cell walls

U. Neugebauer U. Neugebauer et al.et al. ChemPhysChem ChemPhysChem 20072007, , 88, 124., 124.

C. Budich C. Budich et alet al. J. Microsc. . J. Microsc. 20072007, (accepted), (accepted)

TERS setupTERS setup

Coarse xy AFM-positioning

Piezo z-focusing

Piezo xy sample stage

Coarse xy sample stage

AFM

IFNF Laser

Spectrometer & CDD

A. Rasmussen, V. Deckert, J. Raman Spectrosc. 2006, 37, 311.

Bottom illumination configuration

TERS tips preparationTERS tips preparation

AgThermal source

Si AFM tip

Low Rate ~ 0.05 nm/s

Diameter ~20 nm

Tip and laser alignmentTip and laser alignment

Direct reflected image

X

PD

Sample: Sample: Synthetic RNA homopolymer Synthetic RNA homopolymer

on micaon mica

Topographical Image

Single RNA strandSingle RNA strand

AFM image Profile

TERS experimentTERS experiment

15s,1mW, @ 530 nm

E. Bailo and V. Deckert, E. Bailo and V. Deckert, Angew. Chem. Int. Ed., Angew. Chem. Int. Ed., 20072007, , accepted.accepted.

Band assignmentBand assignment

CCC

CC

CCCCCCCC

Enhancement factorEnhancement factor

~ 60 bases

Laser spot

~ 1 µm

Ag Tip

Radius ~ 20 nm

~ 3000 bases

50 Fold

Signal-to-noise ratio

200:1x

104

Enhancement factor at least

Spectral fluctuationsSpectral fluctuationsAg

Particle

Band fluctuations:Band fluctuations:adenine-silver complexesadenine-silver complexes

H. Watanabe et al., Phys. Rev. B 2004, 69, 155418.

Predicted Raman spectra:

Vibrational frequencies

and

Raman intensities

change for each isomer

Ad-N1

Ad-N3

Ad-N7

Ad-N10

SERS

TERS

Adenine

Band identificationBand identification

J. Florian et al. J. Phys. Chem. 1996, 100, 5578.

TERS on TERS on poly(cytosine)poly(cytosine)

RamanRaman

on cytosineon cytosineVibrational modesVibrational modes

801801 803803

10451045 10451045

10971097 11061106

11661166 11461146

12481248 12451245

12791279 12731273

13801380 13791379

14531453 14581458

14881488 14901490

15491549 15311531

16021602 16061606

Ag particle

TERS and DNA sequencing TERS and DNA sequencing

Summary and OutlookSummary and Outlook

Single RNA strands detectionSingle RNA strands detection Approx. 60 nucleobasesApprox. 60 nucleobases SNR ~ 200SNR ~ 200

Next step Next step Natural DNA or RNA sequencingNatural DNA or RNA sequencing

AcknowledgementAcknowledgement

Volker DeckertVolker Deckert

Christian Budich, Marc RichterChristian Budich, Marc Richter

H. Herzog, E. Pulvermacher, B. GosciniakH. Herzog, E. Pulvermacher, B. Gosciniak

Tip shapeTip shape

TIP

Sample