Principles of Fluorescence Spectroscopy

176 FREQUENCY-DOMAIN LIFETIME MEASUREMENTS
Alternative technologies are available to obtain FD
measurements at frequencies above 200 MHz. The need for
a light modulator can be eliminated by using the harmonic
frequency content of a laser pulse train. Suppose the light
source consists of a mode-locked argon ion laser and a cavity-dumped
ps dye laser. This source provides 5-ps pulses
with a repetition rate near 4 MHz. In the frequency domain
this source is intrinsically modulated to many gigahertz, as
shown by the schematic Fourier transform in Figure 5.23
(lower panel). The idea of using the harmonic content of a
pulse train was originally proposed for pulsed lasers 99 and
later for synchrotron radiation. 100–102 Pulse sources provide
intrinsically modulated excitation at each integer multiple
of the repetition rate, up to about the reciprocal of the pulse
width. 103–104 For a ps dye laser the 4-MHz pulse train can be
used for frequency domain measurements at 4, 8, 12, 16
MHz, etc. These harmonics extend to GHz frequencies,
which are higher than the upper frequency limit of most
detectors.
There are significant advantages in using the pulses
from a ps laser. The cavity-dumped output of dye lasers
is rather easy to frequency double because of the high
peak power. Frequency doubling provides wavelengths
for excitation of proteins and other extrinsic probes absorbing
in the UV. Importantly, when using a ps dye laser
source it is no longer necessary to use an electrooptic modulator
and nearly crossed polarizers, which results in a significant
attenuation of the incident light. There appears
to be no detectable increase in noise up to 10 GHz, suggesting
that there is no multiplication of phase noise at the higher
harmonics.
The second obstacle to higher frequency measurements
was the detector. The PMT in the 200-MHz instrument
(Figure 5.7) is replaced with a microchannel plate (MCP)
PMT. 105–108 These devices are 10- to 20-fold faster than a
standard PMT, and hence a multi-gigahertz bandwidth was
expected. As presently designed, the MCP PMTs do not
allow internal cross-correlation, which is essential for an
adequate signal-to-noise ratio. This problem was circumvented
by designing an external mixing circuit, 105–106 which
allows cross-correlation and preserves both the phase and
the modulation data. The basic idea is analogous to Figure
5.10, except that mixing with the low-frequency signal is
accomplished after the signal exits the MCP PMT. External
cross-correlation was found to perform well without any
noticeable increase in noise.
What range of frequencies can be expected with a
pulsed-laser light source and an MCP PMT detector? For
Lorentzian-shaped pulses the harmonic content decreases
to half the low-frequency intensity at a frequency ω 2 = 2 ln
2/∆t, where ∆t is the pulse width. 104 For 5-ps pulses the harmonics
extend to 280 GHz, higher than the upper frequency
limit of any available detector. Hence for the foreseeable
future the measurements will be limited by the detector.
The upper frequency of a detector is limited by the
pulse width due to a single photoelectron, or equivalently
the transit time spread. Hence, one expects the highest modulation
frequencies to be measurable with MCP PMTs that
have the smallest transit time spread (Table 4.1). The relative
power at various frequencies can be measured with a
spectrum analyzer. This was done for several PMTs using a
ps pulse train with its high harmonic content as the light
source. These results show that the side-window R928 is
most useful below 200 MHz (Figure 5.24), and cannot be
used for measurements much above about 400 MHz. The
R-1564U is a 6-micron MCP PMT, and shows a useful
response to 2 GHz. This PMT was used in the first 2-GHz
instrument. 105
To obtain frequencies above 2 GHz it was necessary to
use a specially designed MCP PMT, the R-2566. The data
in Figure 5.25 are for the 6-micron version of the R-2566,
which provides measurable power to 10 GHz, and allowed
construction of a 10-GHz FD instrument. 106 This MCP
PMT possesses a grid between the microchannel plates and
the anode, which serves to decrease the width of the photoelectron
pulses. In the frequency domain the upper limit of
the detector is determined by the reciprocal of the pulse
width. In TCSPC the time resolution is determined by the
rise time of the PMT, and the overall pulse width is less
important.
Figure 5.24. Measured frequency-response of several PMTs, and a
fast photodiode (PD). Data from [107] and [108], and technical literature
from Hamamatsu Inc.