a landmark-oriented algorithm to generate the
views, and then applies a mean-intensity algorithm to generate a fusion of the views.
Optimizations for large-area live cell imaging include CMOS detectors rather than point
detectors and the use of a water-immersion
objective lens. LSFM’s value is in observing
samples like developing embryos—at subcellu-lar resolution—over the course of hours or days.
Because it’s essentially a variation of epi-fluo-rescence imaging, LSFM has been successfully
combined with super-resolution techniques
The fusion of technologies
The optical microscope has long been the starting
point for much more specialized instruments.
Technologies such confocal imaging, stimulated
depletion emission, hyperspectral optics and
x-ray imaging all begin, fundamentally, with a
standard upright or inverted optical lightpath.
As with many hybrids, however, the marriage
of methods leaves room for improvements.
Modularity, such as that seen in Olympus
America Inc.’s (Center Valley, Pa.) IX series of
inverted microscopes, has allowed for several
subsystems to be integrated in the lightpath
of the instrument, without interfering with
research workflows or benchtop space. These
subsystems can be exchanged at will, letting one
instrument become as useful as several.
But other systems, such as dedicated Raman
microscopes, can benefit from further, more permanent, integration. Traditionally, Raman instruments were built from an inverted microscope.
The addition of an excitation laser, light detector,
filters and a spectrometer or monochromator
allows the acquisition of spectral light, which in
turn supplies important chemical information
that eludes conventional microscopy.
Early Raman microscopes were bulky, because
these systems were separately packaged and
linked with fiber-optic couplings. More recent
Raman instruments, such as the IDRaman micro
introduced in September 2013 by Ocean Optics,
Dunedin, Fla., have adopted new focusing technology that gives it a compact 4-in by 14-in by
11-in footprint. The instrument’s OneFocus fea-
ture optimizes the instrument for Raman sam-
pling using the same focal plane for collecting
images and Raman signals. This is an improve-
ment over the sampling approach taken by sys-
tems based on traditional inverted microscopes
and fiber-optic couplings. The ability to focus for
optimal Raman sampling while viewing a quality
image of the sample simplifies the often tedious
and inexact process of acquiring data from a
specific structure or location on
a sample. OneFocus also
enhances data collection
for applications where only
a single layer of material
is applied to the surface,
such as graphene or surface
enhanced Raman spectros-
The IDRaman micro is available with either
a 532-nm or a 785-nm laser for excitation and
offers the option for a high-resolution detector
with four wavenumber resolution acquiring
data from 200 to 2,000 wavenumbers, or a
wide-range system with eight wavenumber resolution acquiring data from 200 to 3,200 wavenumbers. Its 3-MP imager uses epi-illumina-tion techniques and interchangeable objectives,
allowing users to adjust spot size and optical
Microscopy companies are making greater
efforts at system integration on a variety of levels,
and individual components reflect this change.
Olympus, for example, began marketing a new
type of image sensor a little more than a year ago
that offers both color and monochrome imaging
in a single unit. Designed for researchers who
previously needed to align two cameras to a
dual port, the system is essentially two complete,
high-performance parfocal and parcentric cameras in one. Geared for the acquisition of both
fluorescence and color brightfield, the DP80 dual
CCD device helps increase functionality on a
single microscope stand.
The basis for this innovation is pixel-shifting
technology that also serves to allow Olympus’
DP73 3-CCD camera to capture three-color
RGB images within a single pixel to improve
resolution. In the case of the DP80 camera, the
shifting pixels offer 12. 5 MP of color brightfield
imaging and 14-bit monochrome imaging for
fluorescence. cellSens software, which performs
post-imaging optimization tools, captures
monochrome data and color information automatically from the two camera sensors in rapid
The future of optical
A greater understanding of the dynamics and
quantum behavior of light has allowed the
development of super-resolution technology.
In addition, advances in mathematics and
algorithms have greatly improved real-time and
Researchers have been working in these
areas for decades. But occasionally, a new tech-
nology appears that takes a unique approach
to improving optical imaging. In 2013, TAG
Optics Inc., Princeton, N.J., introduced the sec-
ond version of its high-speed varifocal optical
device, the TAGLens 2.0. The lens was hailed by
R&D Magazine’s R&D 100 Awards judges and
others for its ability to offer the fastest adaptive
lens speeds on the market. How it does this and
what sets TAGLens apart as an innovative tech-
nology: The device is designed and optimized to
harness small density changes in fluids caused
by sound. The lens uses these changes in density
variations to change the index of refraction,
shaping light as it goes through the device.
According to TAG Optics’ CEO Christian
Theriault, acoustical light-shaping technology
permits scanning speeds 1,000 times faster than
any other existing adaptive lens technology.
The underlying technology was invented at
Princeton Univ. in 2007. A sinusoidal electronic
input in the radio frequency (RF) range creates
a standing sound wave, which drives piezoelec-
tric element vibrations. The TAGLens 2.0’s focal
length and effective aperture are controlled elec-
tronically by adjusting the amplitude and the
frequency of the RF driving signal. Essentially,
says Theriault, this allows the device to feature
multiple effective apertures and the ability to be
operated with multiple frequencies.
Speed improvements are translated into faster scanning routines for 3-D volumes or highly
profiled surfaces, which now test the limits of
high-throughput optical scanning instruments,
which often can’t perform real-time routines
because of the time needed to change focal position or control the depth-of-field independently
of the magnification.
on its head,
using acoustical light-shaping technology to
allow a 1,000-fold improvement in scanning
speeds. Image: TAG Optics Inc.