Selective Two-photon Excitation by Parametrically Shaped Laser

Pulses after an Optical Fiber

Albrecht Lindinger

Institut für Experimentalphysik, Freie Universität Berlin, Arnimallee 14, D-14195 Berlin, Germany

Keywords: Pulse Shaping, Optical Fibers, Multi-photon Processes.

Abstract: Laser pulse shaping through optical fibers is reported for applications on two-photon processes in dye

molecules. The presented method utilizes pre-compensation of the optical fiber properties by analytical

pulse shaping in order to receive specific parametric pulse forms after the fiber including polarization

modulation. Particularly phase-tailored pulse shapes at the distal fiber end are employed for two-photon

fluorescence of dyes in a liquid environment in order to improve the contrast between dye markers having

similar excitation spectra. This will lead to new endoscopic imaging applications with an increased

fluorescence contrast.

1 INTRODUCTION

Coherent control of photo-induced molecular

processes by shaped laser pulses has attained

considerable success in recent years. It became most

exciting when self-learning feedback loop

algorithms were employed where tailored laser

pulses can be generated, which drive the induced

processes at a maximum yield along desired paths

(Judson and Rabitz, 1992); (Brixner and Gerber,

2003). An important issue in this regard is the

information coded in the optimized laser pulse shape

which supplies insight about the underlying

processes (Schäfer-Bung et al., 2004).

In the last years polarization pulse shaping was

explored in order to consider the vectorial character

of the light field (Brixner and Gerber, 2001);

(Polachek et al., 2006). Novel pulse shaper schemes

for simultaneous phase, amplitude, and polarization

pulse control were designed, and a parametric sub

pulse encoding was developed (Weise and

Lindinger, 2010). In this approach, the physically

intuitive parameters energies, distances, and chirps,

as well as the states of polarization of the sub pulses

can be controlled. This yields new perspectives of

adding the polarization and hence utilizing all

properties of the light field in the pulse modulation.

Recently, pulse shaping methods were

increasingly used in life sciences in order to

investigate biologically relevant systems (Herek et

al., 2002). Promising applications are imaging using

multi-photon microscopy, spectroscopy, and

photodynamic therapy (Myaing et al., 2006);

(König, 2000); (Tsen et al., 2007). Laser pulse

shaping was moreover applied to multi-photon

excited fluorescence where interference effects are

utilized (Meshulach and Silberberg, 1998). This

enables to achieve narrow band transitions and can

be used to selectively excite different biologically

relevant fluorophores with close lying excitation

bands (Lozovoy et al., 2002); (Ogilvie et al., 2006).

In order to operate in biologically relevant

environments, in vivo applications are desired were

the light is guided to the place of interest. This can

be realized by transmission of light through optical

fibers for endoscopic applications, which is a novel

topic of large relevance e.g. for medical treatments.

Yet, it is challenging to transfer femtosecond laser

pulses through fibers due to the occurring distortion

of the intense light pulses by linear and nonlinear

optical fiber properties (Agrawal, 2001). Successful

guidance of ultrashort laser pulses would however

enable novel endoscopic imaging and therapeutic

feasibilities.

In this contribution, laser pulse shaping will be

combined with optical fibers and biologically

relevant multi photon spectroscopy. Tailored laser

pulses are guided through hollow core photonic

crystal fibers, and predetermined parametrically

shaped laser pulses in phase, amplitude and

polarization are achieved after the fiber by

Lindinger A..

Selective Two-photon Excitation by Parametrically Shaped Laser Pulses after an Optical Fiber.

DOI: 10.5220/0004337400190024

In Proceedings of the International Conference on Photonics, Optics and Laser Technology (PHOTOPTICS-2013), pages 19-24

ISBN: 978-989-8565-44-0

 2013 SCITEPRESS (Science and Technology Publications, Lda.)

considering the linear, nonlinear, and polarization

fiber properties. This enables to steer photoinduced

processes by utilizing these pulses for multi-photon

excitation in molecular systems. The application of

phase-tailored pulses for imaging contrast

enhancement is finally demonstrated for dye

molecules in a liquid after transmitting a hollow core

photonic crystal fiber.

2 EXPERIMENT

The experimental setup consists of a femtosecond

laser system (Mira oscillator, RegA amplifier,

Coherent Inc.) which delivers 60 fs pulses at 805 nm

central wavelength with a repetition rate of 76 MHz

for the oscillator and 286 kHz for the amplifier.

Pulse shaping is performed by a spatial light

modulator (SLM640, CRi) placed in the Fourier

plane of a 4f-setup for the dye experiments and

combined with another modulator (SLM256, CRi)

and a polarizer in between for the polarization

modulated pulses. The center pixels of the pulse

modulator are aligned to the central wavelength of

805 nm. A wave plate is placed in the beam after the

pulse shaper in order to align the light polarization

parallel to the optical axes of the fiber. The pulses

are subsequently coupled into a hollow core

photonic crystal fiber (HC-800-1, NKT Photonics).

The core of this fiber measures 9.2 μm for the short

axis and 9.5 μm for the long axis. The transmission

window is centered at 830 nm and about 70 nm

wide. The zero dispersion wavelength is located at

805 nm and the third order Taylor term of the

dispersion function dominates for these pulses. Laser

pulses utilized in this contribution exhibit negligible

non-linear effects in the fiber. A telescope is used to

modify the beam diameter for adequate coupling

into the fiber. After the fiber, the laser beam is

widened by a telescope for the dye experiment and

focused into a cuvette containing rhodamine B

(2.5·10

-4

molar) and coumarin 1 (7.5·10

-3

molar)

solved in ethanol.

Since the exact spectral phase of the laser pulses

at the cuvette is crucial for the measurements, its

distortion by the optical elements is compensated by

writing the corresponding phase retardances on the

pulse modulator. The fluorescence light is collected

by two lenses and then detected by a fiber

spectrometer. The fluorescence of the two dyes is

spectrally well separated, which enables to record

the excitation efficiency by integrating from 410 nm

to 550 nm for coumarin 1 and 560 nm to 700 nm for

rhodamine B.

The shaped pulses are detected after the fiber by

using a time resolved ellipsometry scheme. It is

based on a sum frequency generation cross

correlation setup using a BBO crystal (Plewicki et

al., 2006). The shaped pulse is convoluted with a

short reference pulse received directly from the

laser. The BBO crystal is polarization sensitive and

selects only the intensity in one polarization

direction. A set of cross correlation traces is

recorded by rotating the polarization of the shaped

pulse using a half-wave plate. For each time step the

instantaneous ellipse of the electrical field can be

determined. This data is presented in a single graph

in which the time-dependent intensity and

polarization state including ellipticity and orientation

are displayed. The ellipticity is defined as the ratio

of the major to minor axis. A three-dimensional

representation of the pulse is calculated based on the

measured data.

3 PULSE SHAPING THROUGH

FIBERS

Ultrashort laser pulses guided through optical fibers

are affected by the intrinsic optical fiber properties

which have influence on the spectral distribution and

the temporal evolution of the transmitted pulses.

Particularly the dispersion, birefringence and

nonlinearity of the fiber have to be considered.

These properties can be determined by utilizing the

pulse shaper. Generally, spatial light modulator

phase values are searched for in order to pre-

compensate the laser pulse such that one receives a

short pulse after the fiber.

Novel hollow core photonic crystal fibers are

utilized here since they are advantageous for guiding

ultrashort pulses due to the light propagation in the

hollow core and hence their minor dispersion and

nonlinearity (Russel, 2003). The asymmetric shape

of the hollow fiber core leads to large birefringence.

This causes a temporal separation of the components

after propagation through the fiber with each

component chirped differently. Therefore, the fiber

can be described by two perpendicular optical axes,

which have different dispersions. These axes are

denoted as fast (f) and slow (s) axes due to the

different group velocities. If light is linearly

polarized along one of these axes, the state of

polarization is not altered by the fiber. The overall

orientation of the polarization shaped pulse is

determined by the orientation of the optical axis of

the fiber. Both sub pulses are still linearly polarized

PHOTOPTICS2013-InternationalConferenceonPhotonics,OpticsandLaserTechnology

and oriented along the respective optical axes after

propagation through the fiber. The temporal

intensity profiles are asymmetric and broadened due

to chromatic dispersion of the fiber. This broadening

and asymmetry, which is characteristic for second

and third order phase functions, can be compensated

for by applying a phase function of the opposite sign

on each pulse in order to produce short sub pulses

after the fiber. These values corresponding to the

respective axes are written as an offset on the

modulator. The delay between the differently

polarized pulse components is attributed to the

difference in the group velocity and can as well be

pre-compensated with the pulse shaper. Finally, the

relative phase between the two pulses is adjusted to

generate a single linearly polarized output pulse

oriented at 45°. Having this compensation

determined, the pulse which is transmitted through

the fiber can be arbitrarily controlled in phase,

amplitude, and polarization.

Figure 1: Block diagram of the parametric pulse shaping

method through optical fibers.

For the birefringent hollow core fiber the

orientation of the output pulse can be changed by

altering the relative amplitudes of the two

polarization components. The shift of the relative

phase between the components results in a change of

the ellipticity (Weise et al., 2010). This procedure

can be extended to pulse sequences consisting of a

variable number of sub-pulses. Any individual sub-

pulse is generated by two perpendicularly polarized

pulses whose energy ratio, relative phase, and delay

are set. Parametric sub-pulse sequences for each

polarization component are produced by following

the method described in ref. (Weber et al., 2005).

Minor side pulses still occur for the employed pulse

shaper setup, yet they can be removed by a more

advanced pulse shaper design (Weise and Lindinger,

2010).

The determined fiber precompensation phase

functions and the fiber phase difference are included

in the calculation of the phase retardances of the

liquid crystal arrays by adding the corresponding

phase values. The required electric fields at the

proximal end of the fiber can be generated with the

appropriate phase retardances of the liquid crystal

arrays. Hence, combining the desired sub-pulse

sequence phase values with the pre-compensation

phase parameters controls the shape of the laser

pulse after the fiber (see Figure 1 for a

comprehensive method description).

Parametric control of sub-pulses in a pulse

sequence delivered by the Mira oscillator is

illustrated for a series of double pulses, where one

parameter of the pulse sequence is independently

controlled (Figure 2). The double pulses are

constructed from three sub pulse components. The

first sub-pulse in these pulse sequences is linearly

polarized parallel to the slow axis of the fiber. The

two other sub pulse components yield the second

sub-pulse. The experimental application of a linear

chirp on the second sub-pulse and the associated

increase in pulse duration is presented in Figure 2(a).

The rotation of a sub-pulse is demonstrated in Figure

2(b). Both sub-pulses are linearly polarized while

the orientation of the second sub-pulse is changed

from 0 to 150°. The control of the ellipticity from

linear to circular is visualized in Figure 2(c) in

which the second sub-pulse is oriented at −45°

relative to the slow axis of the fiber. These data

prove the good agreement of the set parameters with

the measured parameters of the shaped pulses.

This procedure is not limited to double pulses

and can be extended to a larger number of sub-

pulses and higher complexity, which is exemplified

in Figure 3 where a triple pulse sequence is depicted

with differently chirped and polarized sub-pulses.

The example shows the potential of laser pulse

shaping after optical fibers which can be employed

for various optical applications. Even nonlinear fiber

properties leading dominantly to self-phase

modulation can be controlled by pre-calculating the

input laser pulse shape with back-propagating the

nonlinear Schrödinger equation (Tsang et al., 2003);

(Pawłowska et al., 2012). In the following, pulse

shaping via fibers applied to two-photon dyes will

be demonstrated for one polarization direction in the

linear regime, but it can in principle be extended to

differing polarization directions and to the nonlinear

regime.

SelectiveTwo-photonExcitationbyParametricallyShapedLaserPulsesafteranOpticalFiber

Figure 2: Measured parametric pulse shapes after the

hollow core fiber where the linear chirp (a), the orientation

(b), and the ellipticity (c) of the second sub-pulse are

varied. An exemplary 3-dimensional representation of an

experimentally recorded double pulse is depicted in (d)

where s and f denote the slow and fast axes, respectively.

Figure 3: Shape of a complex laser pulse after the fiber.

The left side displays a 3-dimensional representation of

the measured laser pulse and the right side shows the time-

dependent intensity, ellipticity, and orientation.

4 PULSE SHAPING FOR

TWO-PHOTON PROCESSES IN

DYES

In two-photon processes the photon interference can

be utilized by tailored laser pulses to achieve a

spectrally narrow two-photon excitation (Meshulach

and Silberberg, 1998). This enables selective

excitation of different dyes with partially

overlapping excitation spectra, which can be applied

to two-photon fluorescence microscopy (Ogilvie et

al., 2006). The effective two-photon field is due to

interference of different components within the laser

spectrum E(



) and has the form









 deEEE

i ))()((

000

)2(

)()()2(





with the phase function (



) and the center

frequency



. If the phase function is antisymmetric

around



, the exponent vanishes and leaves for

(2)



) the same result as a transform limited pulse

with a flat spectral phase (Meshulach and

Silberberg, 1998).

In the present contribution sinus phase and third

order phase are employed as antisymmetric phase

functions. They exhibit constructive interference at

the center frequency and partially destructive

interference at other frequencies (see Figure 4). With

the two photon spectrum and the two-photon cross

sections of the involved dyes it is possible to

simulate the excitation frequency dependent relative

fluorescence intensity I of each dye. This permits to

calculate the frequency dependent contrast (I

rhB

cou1

)/(I

rhB

cou1

) between the examined dyes

rhodamine B (rhB) and coumarin 1 (cou1).

For the experiment it is very important to

precisely control the spectral phase after the fiber by

adjusting the pulse shaper settings and carefully

PHOTOPTICS2013-InternationalConferenceonPhotonics,OpticsandLaserTechnology

detecting the pulse shape since the measurements are

phase sensitive. The measurements were performed

Figure 4: Simulated two-photon field for two

antisymmetric phase functions around 815 nm. (a) shows a

third order phase function and (b) a sinus phase function.

Both two-photon spectra exhibit a sharp peak at the point

of antisymmetry at 815 nm which allows selective

excitation.

with amplified laser pulses since the non-linear

excitation requires substantial pulse energy. The

obtained well separated fluorescence spectra of the

two dyes enable to record the specific excitation

efficiencies by integrating the spectral intensity from

410 nm to 550 nm for coumarin 1 and from 560 nm

to 700 nm for rhodamine B.

In Figure 5 frequency scans of the antisymmetric

center of the phase functions were performed after

the fiber for the sinus and third order phases. The

obtained contrast is plotted and shows a good

agreement with the simulation results. Higher

integrated rhodamine B fluorescence compared to

coumarin 1 leads to a contrast larger than zero. For

comparison, the contrast obtained for a transform-

limited pulse is depicted as a horizontal grey line.

The maximal contrast difference is about 0.4 in both

cases which allows a clear separation of the dyes.

The maximal contrast can slightly be increased by

adding higher Taylor terms to the phase functions

since this improves the condition of having

Figure 5: Frequency scans of the point of phase

antisymmetry recording the contrast of the two-photon

excited fluorescence of rhodamine B and coumarin 1. The

intensity is displayed in percentage of a short pulse. A

third order phase of 2·10

is used in (a) and a sinus

function with an amplitude of 25 rad and a wavenumber

of 0.0736 nm

-1

is applied in (b). The experimental data are

in good agreement with the simulations.

constructive interference on one side of the spectrum

and destructive interference on the other side. These

phase scans on these typical dye markers

demonstrate the perspective of pulse shaping after

optical fibers for spectroscopic multi-photon

applications.

5 CONCLUSIONS

Applications of shaped laser pulses via optical fibers

were presented for two-photon processes in dye

molecules. The explained phase, amplitude, and

polarization forming method employs pre-

compensation of the optical fiber properties by

analytical pulse shaping to obtain tailored parametric

pulse forms at the distal fiber end. In a parametric

SelectiveTwo-photonExcitationbyParametricallyShapedLaserPulsesafteranOpticalFiber

sub pulse encoding, the physically intuitive sub

pulse parameters including the polarization state

were individually controlled.

Moreover, anti-symmetrically phase-shaped

pulses after the fiber were applied for two-photon

fluorescence of dyes in liquids to enhance the

contrast between different dyes with similar

excitation spectra. The received experimental results

were found to be in good agreement with the

conducted theoretical simulations which

demonstrates the precision and reliability of the

presented method. This novel technique will have

perspectives in endoscopic imaging applications by

yielding an increased fluorescence contrast.

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PHOTOPTICS2013-InternationalConferenceonPhotonics,OpticsandLaserTechnology