IEEE JOURNAL OF SELECTED TOPICS IN QUANTUM ELECTRONICS, VOL. 25, NO. 1, JANUARY/FEBRUARY 2019

6801408

Scan-Less, Kilo-Pixel, Line-Field Confocal
Phase Imaging with Spectrally Encoded
Dual-Comb Microscopy
Eiji Hase , Takeo Minamikawa , Shuji Miyamoto, Ryuji Ichikawa, Yi-Da Hsieh,
Yasuhiro Mizutani, Tetsuo Iwata, Hirotsugu Yamamoto, and Takeshi Yasui

Abstract—Confocal laser microscopy (CLM) is a powerful toolin life science research and industrial inspection, and its image acquisition rate is boosted by scan-less imaging techniques. However,
optical-intensity-based image contrast in CLM makes it difficult
to visualize transparent non-fluorescent objects or reflective objects with nanometer unevenness. In this paper, we introduce an
optical frequency comb (OFC) to scan-less CLM to give opticalphase-based image contrast. One-dimensional (1-D) image pixels
of a sample are separately encoded onto OFC modes via 1-D spectral encoding by using OFC as an optical carrier of amplitude andphase with a vast number of discrete frequency channels. Then,
Manuscript received April 6, 2018; revised August 16, 2018 and October24, 2018; accepted October 25, 2018. Date of publication November 9, 2018;
date of current version November 20, 2018. This work was supported by theJapanese Science and Technology Agency (JST) under Grant JPMJER1304
for Exploratory Research for Advanced Technology (ERATO) MINOSHIMAIntelligent Optical Synthesizer (IOS) Project. (Corresponding author: Takeshi
Yasui.)
E. Hase is with Research and Utilization Division, Japan Synchrotron Radiation Research Institute, Hyogo 679-5198, Japan, and also with JST, ERATO,MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima 770-8506,
Japan (e-mail:,hase@spring8.or.jp).
T. Minamikawa, T. Iwata, and T. Yasui are with the Graduate School ofTechnology, Industrial and Social Sciences, Tokushima University, Tokushima
770-8506, Japan, and also with JST, ERATO, MINOSHIMA Intelligent Optical
Synthesizer Project, Tokushima 770-8506, Japan (e-mail:,minamikawa.takeo@
tokushima-u.ac.jp; iwata@tokushima-u.ac.jp; yasui.takeshi@tokushima-u.
ac.jp).
S. Miyamoto was with the Graduate School of Advanced Technology and
Science, Tokushima University, Tokushima 770-8506, Japan, and also with
JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima
770-8506, Japan. He is now with SCREEN Semiconductor Solutions Company
Limited, Shiga 520-2323, Japan (e-mail:,shuujimiyamoto@gmail.com).
R. Ichikawa was with the Graduate School of Advanced Technology and
Science, Tokushima University, Tokushima 770-8506, Japan, and also with JST,
ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima 7708506, Japan. He is now with Tsubakimoto Chain Corporation, Kyoto 610-0380,
Japan (e-mail:,richikawa4504@gmail.com).
Y.-D. Hsieh was with the Graduate School of Technology, Industrial and Social Sciences,Tokushima University, Tokushima 770-8506, Japan, and also with
JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima
770-8506, Japan. He is now with MiZ Company Limited, Kanagawa 247-0056,
Japan (e-mail:,heeida@gmail.com).
Y. Mizutani is with the Graduate School of Engineering, Osaka University,
Osaka 565-0871, Japan, and also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima 770-8506, Japan (e-mail:, mizutani@mech.eng.osaka-u.ac.jp).
H. Yamamoto is with the Center for Optical Research and Education, Utsunomiya University, Tochigi 321-8585, Japan, and also with JST, ERATO,MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima 770-8506,
Japan (e-mail:,hirotsugu@yamamotolab.science).
Color versions of one or more of the figures in this paper are available online
at http://ieeexplore.ieee.org.
Digital Object Identifier 10.1109/JSTQE.2018.2879017

line-field confocal information of amplitude and phase are decodedfrom mode-resolved OFC amplitude and phase spectra obtainedby dual-comb spectroscopy. The proposed confocal phase imaging
will further expand the applications of CLM.
Index Terms—Biomedical imaging, optical microscopy, opticalinterferometry.

I. INTRODUCTIONPTICAL microscopy has been widely used as a powerful tool in life science research and industrial inspection.
Therefore, the development of optical microscopy has been aprincipal driving force in these fields. Among various types of
optical microscopy, confocal laser microscopy (CLM) [1], [2]has attracted attention due to its two-dimensional (2D) or threedimensional (3D) imaging capability with confocality, enablinga depth resolution of wavelength order and stray light elimination. Since the confocality is achieved by a conjugate relationship among a light-source pinhole, a focal point, and a detection
pinhole, mechanical scanning of the focal spot is necessary to acquire the image. However, this mechanical scanning hampers the
rapid image acquisition and the robustness to surrounding external disturbance, such as air turbulence or mechanical vibration.
One promising approach to achieve confocal one-dimensional(1D) imaging without the need for mechanical scanning is aspectral encoding method, namely 1D-SE-CLM [3]–[5]. In 1DSE-CLM, 1D image of a sample is encoded onto a broad optical
spectrum based on the dimensional conversion between wavelength and space while giving the confocality, and then thesample image was decoded from the image-encoded spectrum.
Such 1D-SE-CLM enables the scan-less confocal 1D imaging
with the rapid image acquisition rate and high robustness against
external disturbances, of which the efficacy is confirmed in medical applications [6], [7] and endoscopic applications [8], [9]. In
the conventional 1D-SE-CLM, the image contrast is obtainedby the optical intensity, which is sensitive to absorption, scattering, or reflection of objects; however, it is difficult to givea high image contrast to transparent non-fluorescent objects orreflective objects with nanometer unevenness. If another image
contrast sensitive to these objects is given in the scan-less CLM,
its range of applications will be further expanded to imaging ofliving cells or nanometer topography of semiconductor objects.
The use of optical phase is an interesting candidate for the
new image contrast in scan-less CLM because it provides in-O1077-260X © 2018 IEEE. Personal use is permitted, but republication/redistribution requires IEEE permission.
See http://www.ieee.org/publications standards/publications/rights/index.html for more information.

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formation on refractive index, optical thickness, or geometricalshape and hence visualizes transparent non-fluorescent objects
or reflective objects with nanometer unevenness. One possible method for the phase contrast is confocal phase-sensitive
approach [10]–[12]. Since the interference occurs just only between a light that comes from the confocal volume and a parallel
reference beam, each spatial mode can act as a virtual pinhole,enabling the confocal amplitude and phase imaging without theneed for confocal pinholes. Its depth resolution was achieved tonanometer based on the optical phase. However, the mechanicalscanning of a focal spot or a sample is still required to obtainthe line image due to the point measurement. Another possible method is an interferometric confocal microscope using anarray of vertical cavity surface emitting lasers (VCSELs) [13].
Although confocal phase imaging could be achieved with an integration times of 100 μs, the lateral resolution is limited by an
off-axis digital holography configuration. Differential confocal
microscopy is a powerful tool for nanometer axial resolution andmillisecond image acquisition in the confocal imaging by converting the signal intensity variation in a linear confocal slope
into the height information [14]–[16]. However, this approachalso needs mechanical scanning of a focal spot or a sample forline imaging.
Comparing with those previous CLMs, 1D-SE-CLM is more
promising approach for a variety of applications from theviewpoint of the non-scanning line imaging. A potential lightsource for the optical-phase-based contrast in 1D-SE-CLM isan optical frequency comb (OFC) [17]–[19]. OFC is composedof a vast number of discrete, regularly spaced optical frequency
modes, and the optical frequency and phase of all OFC modes
are secured to a frequency standard by its inherent mode-locking
nature and active laser control. Furthermore, current state-ofthe-art dual-comb spectroscopy, namely DCS, enables us torapidly, precisely, and accurately acquire the mode-resolved
OFC spectra of amplitude and phase without the need of any mechanical scanning [20]–[23]. Therefore, a combination of OFC
with DCS gives the optical-phase-based image contrast to 1DSE-CLM as well as the optical-intensity-based image contrast;
we call it 1D spectrally encoded dual-comb microscopy or 1DSE-DCM. In 1D-SE-DCM, after discretely encoding 1D image
pixels of a sample on each OFC mode by a 1D spectral disperser,
1D confocal images of amplitude and phase are respectively decoded from mode-resolved OFC spectra of amplitude and phase
based on the one-to-one correspondence between image pixels
and OFC modes. Sufficient number of OFC modes enables
kilo-pixel 1D imaging even with narrow spectral bandwidth.
Furthermore, the optical-phase-based image contrast provides
information on refractive index, optical thickness, or geometrical shape, enabling us to visualize transparent non-fluorescentobjects or reflective objects with nanometer unevenness unlike
the conventional intensity-based measurement. In this paper,
we demonstrated a proof-of-principle experiment of scan-less,kilo-pixel, line-field confocal phase imaging by 1D-SE-DCM.
II. PRINCIPLE
Fig. 1 shows a principle of operation. After passing througha light-source pinhole, each comb mode of an input OFC is lin-

Fig. 1.

Principle of operation of the proposed method.

Fig. 2.

Experimental setup for scan-less confocal phase imaging.

early dispersed at different angles depending on the wavelength
by a 1D spectral disperser, namely wavelength/1D-space conversion. Then, spatially dispersed OFC modes are line-focused
as a 1D array of focal spots onto a sample with an objective lens.
Optical characteristics of a sample are encoded onto optical amplitude and phase of OFC modes when spatially dispersed OFC
modes were reflected by a sample. Then, those OFC modes are
spatially overlapped again by an inverse propagation of the same
optics, namely 1D-space/wavelength conversion. The confocality is imposed onto the image-encoded OFC by passing througha detection pinhole. Since the mode-resolved OFC spectra ofamplitude and phase reflect the 1D distribution of optical characteristics in the sample, they were acquired by DCS. Finally, 1D
images of amplitude and phase are respectively reconstructedfrom the mode-resolved OFC spectra of amplitude and phase
based on the one-to-one correspondence between image pixels
and OFC modes.
III. EXPERIMENTAL SETUP
Fig. 2 shows a schematic diagram of the experimental setup.
A femtosecond Er-fiber OFC laser (Menlo Systems, GmbH,
Martinsried, Germany, FC1500-250, center wavelength λc =1550 nm, spectral bandwidth Δλ = 70 nm, mean power P =330 mW, repetition frequency fr ep1 = 250,001,484 Hz, carrierenvelope-offset frequency fceo1 = 20,000,000 Hz) was used asa signal OFC for 1D spectral encoding. frep1 and fceo1 werephase-locked to a rubidium frequency standard (Stanford Research Systems, Inc., Sunnyvale, CA, USA, FS725, accuracy =5 × 10−11 , instability = 2 × 10−11 at 1 s) by a laser control

HASE et al.: SCAN-LESS, KILO-PIXEL, LINE-FIELD CONFOCAL PHASE IMAGING WITH SPECTRALLY ENCODED DUAL-COMB MICROSCOPY

system. After passing through an optical bandpass filter (BPF1,
passband = 1538 – 1562 nm) and a beam splitter (BS), the signal OFC light (mean power P = 30 mW) was diffracted by adiffraction grating (G, Spectrogon AB, Täby, Sweden, PC 1200
30 × 30 × 6, groove density: 1200 grooves/mm, efficiency:90%). The resulting 1D spectrograph of OFC modes was relayed and then focused onto the sample along the x-direction bya pair of lenses (Lens1 and Lens2, focal length: 150 mm) andthe objective lens (OL, dry, numerical aperture: 0.25, working
distance: 5.5 mm, focal length: 16.6 mm). After the reflection
by a sample, the signal OFC light inversely propagated the same
optics for the spatial overlapping of each OFC mode, resulting
in encoding of 1D image on the signal OFC spectrum. After
reflecting by BS and passing through a detection pinhole (PH;diameter = 10 μm) equipped with a pair of lenses (Lens3 andLens4; focal length = 16.6 mm), the signal OFC light was
fed into the DCS system. As mentioned above, although thepresent method has the confocality thanks to the interferencewith a parallel reference beam, such “soft pinhole” has less sharp
confocality than the physical pinhole or “hard pinhole”. In thepresent experiment, we used a confocal pinhole with the interferometric configuration to benefit from both “soft ” and “hard”
pinholes.
We used another femtosecond Er-fiber OFC laser (Menlo
Systems, GmbH, Martinsried, Germany, FC1500-250, center
wavelength = 1550 nm, spectral bandwidth Δλ = 70 nm,
mean power P = 330 mW, fr ep2 = 249,998,918 Hz, Δfr ep =fr ep1 − fr ep2 = 2,666 Hz, fceo2 = 20,000,000 Hz) for a local
OFC. fr ep2 and fceo2 were phase-locked to the same frequency
standard by another laser control system. After passing throughanother optical bandpass filter (BPF2, passband = 1538 –1562 nm), the local OFC light was spatially overlapped withthe signal OFC light by a polarization beam splitter (PBS1).
The interferogram was detected by a combination of a halfwave plate (HWP), another polarization beam splitter (PBS2),
and a balanced detector (BD, Newport Corp., Irvine, CA, USA,1617-AC-FS, wavelength: 900-1700 nm, bandwidth: 40 kHz to
800 MHz). We acquired 10 consecutive interferograms with atime window of 40 ns at a sampling interval of 40 fs by a digitizer (Avaldata Corp., Tokyo, Japan, APX-500/ADM-214EX,
sampling rate = 250,000,000 samples/s, number of sampling
points = 937,735, resolution = 14 bit). The mode-resolved amplitude and phase spectra of the signal OFC were obtained by
Fourier transform of the acquired interferogram, and then wereused to decode the 1D amplitude and phase image of the sample. To obtain 2D image of the sample, the sample position
was mechanically scanned along the y-direction perpendicular
to 1D array of focal spots by a stepping-motor-driven translation
stage.
We used a commercialized 1951 USAF resolution test chartwith a negative pattern (Edmond Optics, Inc., Barrington, NJ,
USA, #38-256) for a sample. Negative patterns were fabricated by vacuum-depositing a durable chromium coating on a
float glass substrate (thickness: 1.5 mm) as a reflective film.
The chromium film caused the 2D distribution of both thereflectivity and the phase change corresponding to the chartpatterns.

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Fig. 3. Mode-resolved (a) amplitude and (b) phase spectra of the no-imageencoded OFC. Mode-resolved (c) amplitude and (d) phase spectra of the imageencoded OFC.

IV. RESULTS
A. Mode-Resolved Spectrum of Amplitude and Phase in
1D-Image-Encoded OFC
We first obtained the mode-resolved amplitude and phase
spectra of the no-image-encoded OFC [Figs. 3(a) and 3(b)] andthe image-encoded OFC [Figs. 3(c) and 3(d)] for the test chart.
The no-image-encoded OFC and the image-encoded OFC wereobtained by irradiating the signal OFC onto the no-pattern coating area and the pattern coating area of the test chart. To obtainsufficient SNR, we accumulated 5000 interferograms and thenperformed Fourier transform. Compared with the amplitude andphase spectra of the no-image-encoded OFC, those of the imageencoded OFC were significantly modulated. The unevenness ofthe spectral envelope in Figs. 3(c) and 3(d) reflects the 1D distribution of the reflectivity and phase change in the test chart,
respectively. In this way, we obtained the mode-resolved spectrum of amplitude and phase in the 1D-image-encoded OFC.
B. Confocal Amplitude Imaging
To obtain 2D confocal amplitude image of the test chart, thesample position was scanned at an interval of 1 μm along they-direction while acquiring the mode-resolved amplitude spectrum. Then, a series of the mode-resolved amplitude spectra
were used for 2D confocal amplitude imaging of the test chart.
Fig. 4(a) shows the confocal amplitude image of the test chart(image size = 456 μm × 150 μm, pixel size = 5000 pixels × 150
pixels), in which the horizontal dimension was decoded fromthe OFC spectrum of about 10 nm spectral range whereas thevertical dimension was obtained by the mechanical scanning.
Several test patterns with the spatial frequency of a few tens
lp/mm was clearly confirmed. The horizontal scratch patterns
were mainly due to the residual timing jitter between the signal
and local OFC lasers. To evaluate the spatial resolution in x-yplane or the lateral resolution δx , δy , we extracted the amplitude
profile along a green line and a blue line in Fig. 4(a). Plots andlines in Figs. 4(b) and 4(c) show differential curves of amplitude profiles for these lines and the curve fitting results with aGaussian function, respectively. When the lateral resolution isdefined as a FWHM of the differential curve, δx and δy were

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IEEE JOURNAL OF SELECTED TOPICS IN QUANTUM ELECTRONICS, VOL. 25, NO. 1, JANUARY/FEBRUARY 2019

Fig. 5. (a) 2-D confocal phase image of a USAF resolution test chart at focus(z = 0 μm) and (b) the corresponding 2-D confocal amplitude image. (c) 3-D
reconstructed image of the test chart calculated from the confocal phase image.

Fig. 4. (a) 2-D confocal amplitude image of a USAF resolution test chart at
focus (z = 0 μm). Differential curve for lateral profiles of amplitude value along(b) x coordinate and (c) y coordinate. (d) 2-D confocal amplitude image of thetest chart at out of focus (z = +120 μm). (e) Depth profiles of amplitude value
with respect to the z position.

determined to be 4.9 ± 0.2 μm and 10.9 ± 1.7 μm, respectively.
Since the field-of-view and the spatial resolution in x-direction
were 456 μm and 4.9 μm, the effective number of resolvable
points was determined to be 93 points.
To confirm the confocality in the confocal amplitude image,
we set the test chart at out of focus (z = +120 μm). Fig. 4(d)shows the corresponding confocal amplitude image. The pattern of the test chart completely disappeared, implying the good
confocality in the proposed 1D-SE-DCM system. For the quantitative evaluation of confocality, we measured the change of
amplitude value when the sample position was incrementally
moved at a step of 2 μm along z axis by a translation stage.
Plots and a line in Fig. 4(e) show the depth profiles of amplitude
value and the curve fitting result with a Gaussian function, respectively. From this depth profile, the depth resolution δz wasdetermined to be 50.0 μm. In this way, the confocal effect was
clearly confirmed in the confocal amplitude image.
C. Confocal Phase Imaging
Next, we decoded a confocal phase image of the test chartfrom the mode-resolved phase spectrum of the image-encoded

OFC. To eliminate the influence of the initial phase in the signal OFC, we subtracted the phase spectrum of the no-imageencoded OFC from that of the image-encoded OFC. Fig. 5(a)shows a confocal phase image of the test chart (image size =190 μm × 100 μm, pixel size = 2084 pixels × 100 pixels)
when we set the chart in focus (z = 0 μm), and Fig. 5(c) shows
the 3D reconstructed image of Fig. 5(a). For comparison, thecorresponding confocal amplitude images is shown in Fig. 5(b).
Test chart pattern clearly appeared in both images. However, theimage contrast mechanism is different between them: the phasedifference for the confocal phase image and the reflectivity difference for the confocal amplitude image between the patterned
region and the surrounding region, which are corresponding
to the reflection-coated region and the uncoated region, respectively. The mean phase difference between them was determined
to be 0.57 ± 0.11 rad by subtracting the mean phase value at thecoated area from that at the non-coated area.
The confocal phase image in Fig. 5(a) enables us to calculate
the thickness of the reflection-coating film. The relative height
H(x,y) is given byλ
1 φ (x, y)λ=
φ (x, y)
(1)
2 2π
4π
where λ is a typical wavelength of OFC and Φ(x, y) is a confocal
phase image. From the mean phase difference of 0.57 ± 0.11 radand λc of 1550 nm, the mean thickness of the reflection-coating
film was calculated to be 70.7 ± 13.3 nm. On the other hand,the specification value of the reflection-coating film is given
to be around 100 nm. The reason for difference between theexperimental value and the specification one is discussed later.
H (x, y) =V. DISCUSSION
We first estimated the theoretical value for the lateral resolution δx , δy of the proposed 1D-SE-DCM. Theoretical δx and δyvalues are given by [2], [24]
f ·λ
W
0.61λ
δy =
NA

δx =

(2)
(3)

HASE et al.: SCAN-LESS, KILO-PIXEL, LINE-FIELD CONFOCAL PHASE IMAGING WITH SPECTRALLY ENCODED DUAL-COMB MICROSCOPY

where f is a focal length of OL ( = 16.6 mm), W is a diameter ofthe input beam waist ( = 8 mm), and NA is a numerical apertureof OL ( = 0.25). From (2) and (3), theoretical δx and δx values
were estimated to be 3.2 μm and 3.8 μm, respectively. On theother hand, experimental δx and δy values were 4.9 ± 0.2 μm
and 10.9 ± 1.7 μm, respectively. The beam shape was horizontally oval at the entrance pupil of the objective lens due to thereflection grating. While the x dimension of the horizontallyoval shape just filled the entrance pupil of the objective lens, they dimension of it did not fill it fully. Difference of effective NA
between the x and y directions is the reason for the large difference of δy between the experimental and the theoretical values.
The better spatial resolution will be achieved for biomedical investigation and application if a higher-NA objective lens can beused. For example, theoretical δx and δy values are estimated tobe 0.64 μm and 0.74 μm by use of a water-immersion, high-NA
objective lens (NA = 1.27, f = 3.33 mm).
We also estimated the theoretical value for the depth resolution δz . Theoretical δz value is given by [2], [24]δz = 0.88

λ√n − n2 − N A2

(4)

where n is the reflective index of the medium between the objective lens and the sample ( = 1). From (4), δz was estimatedto be 43.0 μm by using NA of 0.25. This theoretical value wasin good agreement with the experimental δz value ( = 50 μm).
The present experimental δz value is simply limited by use of
low-NA objective lens. The simple way to improve the depth
resolution is use of high-NA objective lens. For example, if awater-immersion, high-NA objective lens (NA = 1.27, n = 1.33)
is used, the depth resolution of 1.46 μm can be achieved. Also,
the confocal gating is another way to improve the depth resolution [10]–[13]; only the light that comes from the multiple
focal points of the signal OFC is interfered with a properly collimated light of the local OFC. No use of the confocal pinhole
will increase both the SNR and the depth of focus.
We next discuss the difference between the experimental
value ( = 70.7 ± 13.3 nm) and the specification one (ࣈ100 nm)
of the reflection-coating film thickness in the test chart. Whenthe surface unevenness is made in the same material, the phasedifference is directly related to the geometrical shape. However,if the surface unevenness is made by different materials, thephase difference depends on the geometrical shape and difference of complex refractive index between different materials.
Since the surface unevenness of the test chart was made by thecoating film and the float glass substrate, it is corresponding
the latter. When the light is normally incident on a sample, thephase shift θ of the reflected light is derived from the following
equations [25], [26]n + iκ − 1 √ iθ
N −1
=
≡ Re
N +1
n + iκ + 1
√22
 εr − 1  2
 = (1 − n) + κ
R =  √2
εr + 1(1 + n) + κ2r̂ =

θ = tan−1−2κn2 + κ2 − 1

(5)
(6)
(7)

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where r̂ is the Fresnel coefficient, N is the complex refractive
index, n and κ are the real and imaginary part of the complex
refractive index, R is the reflectivity, and εr is the dielectric
constant. By substituting the complex refractive indices of thechromium (n = 4.18, κ = 4.90) [25] and the float glass (n =1.51, κ = 0) [27] into those equations, difference of complex
refractive index between the coating film and the float glass
substrate results in the phase shift of −0.24 rad at the reflection.
By correcting (1) with this phase shift, the final thickness of thereflection-coating film was determined to be 100.3 ± 13.3 nm,
which was in good agreement with its specification value. Thisconsistency clearly indicated a high potential of 1D-SE-DCM
for nanometer surface topography.
It is important to clarify difference between the present 1DSE-DCM and the previous 1D-SE-CLM [3], [4]. One important
advantage of the present 1D-SE-DCM is the optical-phase-based
image contrast although the scan-less confocal imaging can beachieved in both the 1D-SE-DCM and previous 1D-SE-CLM.
Importance and usefulness of the optical-phase-based image
contrast were described in the introduction and were demonstrated in the nanometer surface topography of the test chart.
More interestingly, if δz is achieved to the phase wrapping period (λ or λ /2), the phase wrapping ambiguity, which is always
an obstacle in phase imaging, can essentially be solved because
the confocality suppresses the phase wrapping due to its depth
selectivity. Fortunately, there is still sufficient space to decreaseδz down to sub-micrometer by use of an oil-immersion high-NA
objective lens. In this case, the combination of confocality andphase imaging, namely confocal phase imaging, will enhancethe depth range from nanometer to micrometer to millimeterscale in surface topographic applications or will make it possible
to visualize thick biological specimens, which introduce phaseshifts of more than 2π, without the phase wrapping ambiguity.
Another advantage is a large number of pixels in the 1D image along the spectral-encoding coordinate or the x coordinate.
Since the signal OFC had a spectral bandwidth Δν of 1.25 THzaround 192 THz, corresponding to 10.1 nm and 1550 nm inwavelength, and fr ep of 250 MHz, the number of pixels wasachieved to 5000. On the other hand, when the similar Δν wasused for 1D-SE-CLM equipped with a commercialized optical
spectrum analyzer (namely, OSA-based 1D-SE-CLM, spectral
resolution ࣈ0.05 nm or 6.24 GHz), the number of pixels remained around 200. If a dispersive Fourier transform (parallel
detection channels = 8, sampling rate of a digitizer = 50 ×
109 sample/s) is applied for 1D-SE-CLM, namely dFT-based
1D-SE-CLM [4], [24], the number of pixels is increased up to1600; however, it still falls short of that in 1D-SE-DCM. If 1DSE-DCM is combined with the precise scanning of line focused
beam along the y-direction, high-definition (HD) or mega-pixel
2D confocal imaging of amplitude and phase will be achieved.
Also, the proposed 1D-SE-DCM has still room to increase thenumber of pixels up to several tens thousands by expanding Δν
and/or decreasing fr ep . Huge number of pixels enables us toexpand the scan-less confocal amplitude and phase imaging to
2D imaging [28] by help of 2D spectral encoding [5], [29], [30].
One may query whether the kilo-pixel is required for 1D
imaging because the present system could not optimize 5000

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pixels due to its limited effective number of resolvable points(= 93). It is challenging to achieve more than 1,000 resolvable points for a commercially available microscopy objective
lens. However, a combination of 8K ultra-high-definition camera with a spinning disk enables to image a 1 mm2 field with alateral resolution of 0.68 μm, corresponding to 1470 resolvable
points [31]. Using this combination, individual activity of a single synapse was clearly visualized from the wide-field calcium
imaging. Such kilo-pixel by kilo-pixel pixel wide-field imaging
will be a powerful tool for biomedical imaging in the future.
However, the dynamic range of signal intensity is limited to
10 bit while the wide-field imaging was performed at a frame
rate of 60 fps. On the other hand, the 1D-SE-DCM has a potential
to expand the dynamic range by use of a wide-dynamic-range
photodetector while maintaining the similar pixels numbers.
Finally, we compare the 1D-image acquisition rate among
1D-SE-DCM, OSA-based 1D-SE-CLM, and dFT-based 1DSE-CLM. The image acquisition rate in 1D-SE-DCM is limited by the measurement rate of DCS, which is equal toΔfr ep (= 2,666 Hz), and the number of signal accumulation.
In the demonstration of this article, sufficient number of signal accumulation (ࣈ5000) leads to relatively slow acquisition
rate ( = 0.53 line/s). However, there is still space to increase
the imaging speed up to Δfr ep without the need for the signal
accumulation by using tightly-locked dual-comb sources [28].
Furthermore, the optical amplification of the image-encoded
OFC by an erbium-doped fiber amplifier (EDFA) will benefit
high SNR in such a single-shot measurement. The image acquisition rate in OSA-based 1D-SE-CLM is limited to arounda few tens Hz due to the mechanical scanning of a diffraction grating in OSA. A combination of superluminecent diodes(SLD) with a fast spectrometer can further boost the image acquisition rate in 1D-SE-CLM up to hundreds of kHz or evenover 1 MHz. However, if a sample has wavelength-dependent
absorption, the broad spectrum of SLD may suffer from such
absorption. Also, chromatic aberration of the objective lens isanother problem. On the other hand, high spectral resolution of
the 1D-SE-DCM enables the sufficient number of pixel eventhough a broadband light source with narrower spectral bandwidth is used. Use of such light source will reduce influenceof the wavelength-dependent absorption and/or the chromaticaberration. dFT-based 1D-SE-CLM can boost the image acquisition rate up to fr ep (typically, a few MHz to several tens
MHz) thanks to the time-stretch approach. In this way, the acquisition rate in 1D-SE-DCM is higher than that in OSA-based
1D-SE-CLM, although it could not reach that of a dFT-based
1D-SE-CLM. However, in actual fact, the maximum acquisition
speed is often limited by the signal level from the sample undermaximum allowable illumination power.
VI. CONCLUSIONS
We successfully introduce a new image contrast based on
the optical phase to scan-less CLM by use of the 1D-imageencoding OFC with DCS. Confocal 1D images of amplitude
and phase in the sample were simultaneously decoded from themode-resolved spectra of amplitude and phase in the 1D-imageencoding OFC. Nanometer surface topography of the test chart

was demonstrated with high precision. If the image quality andimage acquisition rate are further improved by reduction of thetiming jitter between dual OFC lasers as well as optimization ofΔν, fr ep , and Δfr ep , the proposed method will be a powerfultool in a variety of applications. Potential applications of the proposed method include the surface topography of semiconductor
objects and non-staining 3D imaging of living cells and tissues.
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Eiji Hase was born in Tokushima, Japan, in 1989.
He received the B.S., M.S., and Ph.D. degrees inengineering from Tokushima University, Tokushima,
Japan, in 2012, 2014, and 2017, respectively.
From 2014 to 2017, he was a Research Associatewith JST, ERATO, MINOSHIMA Intelligent Optical
Synthesizer Project, Tokushima, Japan. Since 2017,
he has been a Research Scientist with the Researchand Utilization Division, Japan Synchrotron Radiation Research Institute, Hyogo, Japan, and a Visiting Associate Professor with the Graduate Schoolof Technology, Industrial and Social Sciences, Tokushima University. His research interests include biomedical imaging and biomechanical analysis using
femtosecond laser and synchrotron X-ray.
Dr. Hase was a recipient of the Optical Society of Japan Best Presentation
Award 2016, Japanese Society for Medical and Biological Engineering Symposium Award 2016, and the International Society for Optical Engineering (SPIE)
BiOS2016 in Photonics West 2016 Student Poster Awards in 2016. He is a
Member of SPIE.

Takeo Minamikawa was born in Ibaraki, Japan, in
1983. He received the B.S., M.S., and Ph.D. degrees
in engineering from Osaka University, Osaka, Japan,
in 2006, 2008, and 2010, respectively.
From 2010 to 2013, he was a Research Fellow under the Research Fellowship for Young Scientists of
the Japan Society for the Promotion of Science. From2013 to 2015, he was an Assistant Professor with the
Department of Pathology and Cell Regulation, Graduate School of Medical Science, Kyoto Prefectural
University of Medicine, Kyoto, Japan. Since 2015,
he has been an Associate Professor with the Department of Mechanical Science, Division of Science and Technology, Graduate School of Technology,
Industrial and Social Sciences, Tokushima University, Tokushima, Japan. He is
also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project,
Tokushima, Japan. He is the author of two book chapters and more than 30
articles, credited with more than 15 inventions apart from holding three patents.
His research interests include Raman microspectroscopy and optical-frequencycomb.
Dr. Minamikawa is the recipient of the Japan Society for Precision Engineering Best Presentation Award 2016, Japanese Society of Pathology Poster
Presentation Award 2015, and the Funai Foundation for Information Technology Research Award 2011. He is a Member of the Optical Society of America,
the Japan Society of Applied Physics, the Optical Society of Japan, the Laser
Society of Japan, the Japan Society of Mechanical Engineers, and the Japan
Society for Precision Engineering.

6801408

Shuji Miyamoto was born in Tokushima, Japan,
in 1992. He received the B.S. and M.S. degrees inmechanical engineering from Tokushima University,
Tokushima, Japan, in 2015 and 2017, respectively.
From 2014 to 2017, he was working on nonlinear
optics under Prof. Takeshi Yasui, Tokushima University, focusing on dual-optical-comb spectroscopy
and scan-less confocal microscope using broadband
light source. Since 2017, he has been an Engineer
with SCREEN Semiconductor Solutions Company
Limited, Shiga, Japan. He is also with JST, ERATO,MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima, Japan.
Mr. Miyamoto was a recipient of the Japanese Society for Medical and Biological Engineering Branch Conference Young Researcher Award in 2016.

Ryuji Ichikawa was born in Kohchi, Japan, in 1990.
He received the B.S. and M.S. degrees in engineering from Tokushima University, Tokushima, Japan, in2013 and 2015, respectively. Since 2015, he has been
an Engineer with the Tsubakimoto Chain Corporation, Kyoto, Japan. He is also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project,
Tokushima, Japan. His research interests include THz
gas spectroscopy and dual THz comb spectroscopy.

Yi-Da Hsieh was born in Kaohsiung, Taiwan, in
1984. He received the B.S. and M.S. degrees inbiomedical imaging and radiological sciences from
National Yang-Ming University, Taiwan, in 2006
and 2008, respectively, and the Ph.D. degree fromthe Graduate School of Engineering Science, Osaka University, Osaka, Japan, in 2014. From 2014
to 2016, he was a Postdoctoral Fellow with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer
Project. Since 2017, he has been an Engineer with
the MiZ Company Limited, Kanagawa, Japan. He is
also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project,
Tokushima, Japan. His areas of research are THz instrumentation and opticalfrequency-comb.

Yasuhiro Mizutani received the B.E. and M.E. degrees in nuclear engineering from Osaka University,
Osaka, Japan, in 1997 and 1999, respectively. He obtained the Ph.D. degree in mechanical engineeringfrom the Tokyo University of Agriculture and Technology, Tokyo, Japan, in 2008.
From 1999 to 2003, he worked as a Researcher
with Panasonic Corporation. Then, from 2003 to
2009, he worked as a Research Associate with the Department of Mechanical Systems Engineering, Tokyo
University of Agriculture and Technology. In 2009,
he joined the University of Tokushima as an Associate Professor. Since 2015,
he has been working as an Associate Professor with Osaka University. He is
also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project,
Tokushima, Japan. He has authored or coauthored over 100 papers in refereed
international journals and conferences in the areas of his research interest.
His research interests include interferometry, polarimetry, 3-D surface measurement, optical trapping, and 3-D lithography. He is a Member of the International Society for Optical Engineering and the Optical Society of America.

6801408

IEEE JOURNAL OF SELECTED TOPICS IN QUANTUM ELECTRONICS, VOL. 25, NO. 1, JANUARY/FEBRUARY 2019

Tetsuo Iwata received the Ph.D. degree in engineering from Osaka University,
Osaka, Japan, in 1984.
From 1984 to 1998, he worked with JASCO Corporation, Japan. He iscurrently a Professor with the Graduate School of Technology, Industrial andSocial Sciences, Tokushima University, Tokushima, Japan. He is also with JST,
ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima,
Japan.
His research interests include fluorescence measurement and spectroscopy.

Hirotsugu Yamamoto received the Ph.D. degree in
information science and technology from the University of Tokyo, Tokyo, Japan, in 2008.
From 1996 to 2014, he worked with Tokushima
University, Tokushima, Japan. He is currently an Associate Professor in the Graduate School of Engineering, Utsunomiya University, Utsunomiya, Japan. He
is also with JST, ERATO, MINOSHIMA Intelligent
Optical Synthesizer Project, Tokushima, Japan.
His research interests include information
photonics.

Takeshi Yasui received the first Ph.D. degree inengineering from the University of Tokushima,
Tokushima, Japan, in 1997, and the second Ph.D.
degree in medical science from the Nara Medical
University, Yagi, Japan, in 2013.
From 1997 to 1999, he worked as a Postdoctoral
Research Fellow with the National Research Laboratory of Metrology, Japan. He was with the Graduate School of Engineering Science, Osaka University
from 1999 to 2010, and was briefly with the University of Bordeaux I in 2007 and 2012, and also with
the University of the Littoral Opal Coast in 2010 as an Invited Professor. He iscurrently a Professor with the Graduate School of Technology, Industrial andSocial Sciences, Tokushima University, in addition to being the Vice Directorof Research Support in the same university since 2016. He is also an Invited
Professor with the Graduate School of Engineering Science, Osaka University,
Toyonaka, Japan. He is also with JST, ERATO, MINOSHIMA Intelligent Optical Synthesizer Project, Tokushima, Japan. His research interests include THz
instrumentation and metrology, second-harmonic-generation microscopy, andoptical frequency comb.
Prof. Yasui is a Member of the Optical Society of America, the International Society for Optical Engineering, the Japan Society of Applied Physics,
the Optical Society of Japan, the Laser Society of Japan, the Japanese Society
for Medical and Biological Engineering, and the Japan Society of MechanicalEngineers. He was a recipient of the Award for the Most Promising Young
Scientist from the Optical Society of Japan in 1998, the Sakamoto Award fromthe Japan Society of Medical Electronics and Biological Engineering in 2006,
the Optics Paper Award from the Japan Society of Applied Physics, the Funai
Award from the Japan Society of Mechanical Engineers in 2009, and the Original Paper Award from the Laser Society of Japan in 2013.