Biomedical Engineering Reference
In-Depth Information
×10
−4
20
(3)
(
χ
R
)
2
(3)
(
χ
NR
)
2
15
(3)
(3)
2Re(
χ
R
)
χ
NR
I
CARS
10
5
0
-5
-800
-600
-400
-200
0
200
400
600
800
δ
(cm
−1
)
FIgurE 13.3
Spectrum of the CARS signal and of its different components for a single Raman-active line.
Efforts to remove the nonresonant background have led to the development of various methods,
polarization-sensitive detection, time-resolved CARS [27], epidetection, but at the expense of an attenu-
ation of the resonant signal [20-22]. This disadvantage was suppressed in a method proposed in Ref.
[24], where an intermediate image Δ
i
is formed by subtracting the image
I
−
at the dip frequency from
the image
I
+
at the peak frequency. The image
I
bg
of the surrounding medium is recorded and the cor-
rected image is obtained by calculating the image ∆
I I
b
/ . However, this method increases the overall
acquisition time since two images must be recorded. Ganikhanov et al. proposed an efficient method
to rapidly acquire the image Δ
I
using frequency-modulated CARS (FM-CARS) [25]. In FM-CARS, the
vibrational band probed is frequency-modulated using a Pockels cell to rapidly switch between two
pump beams at the peak and dip frequencies of the CARS spectrum. Highly sensitive detection of the
resulting amplitude-modulated CARS signal is then performed after amplitude demodulation with a
lock-in amplifier. Sensitivity improvements by three orders of magnitude over conventional CARS have
been reported. Simultaneous sensing of two Raman bands was also proposed as a means to suppress
the nonresonant background [26], where this is achieved by calibrating the nonresonant background
ratio between the two CARS signals with a sample that exhibits no vibrational resonance within these
two bands. Contrast improvements by a factor of three were reported. Similar to dual CARS, differen-
tial CARS uses the probing of two Raman bands to obtain background-free images. This is efficiently
achieved using a single detector and a single pump laser however [28]. A replica of the pump-Stokes
pulse train is delayed by half the repetition period and its beat frequency is adjusted by mean of glass
dispersion. The differential-CARS (D-CARS) image between these two Raman bands is then extracted
after lock-in amplification.
While HG and MEF can readily be recorded with the same femtosecond pulsed laser, CARS neces-
sitates two tightly synchronized picosecond pulsed lasers at different frequencies. Picosecond pulses are
preferable to femtosecond pulses for generating CARS because the spectral width of a picosecond pulse
matches that of a Raman line. The energy is thus concentrated on the Raman line width, which helps
improve the resonant to nonresonant background ratio. Precise temporal synchronization between the
pump and Stokes pulses must be attained to create the multiphoton effect and can be achieved with
adjustable delay lines.
