June 1, 2015 / Vol. 40, No.

11 / OPTICS LETTERS 2457

Scanning balanced-path homodyne I/Q-interferometer

scheme and its applications
Seang Hor Eang,1 Seunghyun Yoon,1 Jun Gyu Park,1 and Kyuman Cho1,2,*
1
Department of Physics, Sogang University, 1 Sinsu-Dong, Mapo-Gu, Seoul 121-742, South Korea
2
Interdisciplinary Program of Integrated Biotechnology, Sogang University, 1Sinsu-Dong, Mapo-Gu, Seoul 121-742, South Korea
*Corresponding author: kcho@sogang.ac.kr
Received March 18, 2015; revised May 2, 2015; accepted May 3, 2015;
posted May 5, 2015 (Doc. ID 236471); published May 19, 2015
The balanced-path scheme of a heterodyne interferometer proposed by Yoon et al. has been applied to the scanning
homodyne I/Q-interferometer. This provides an 11-dB improvement on common vibration rejection over the hetero-
dyne scheme and, thereby, allows high sensitivity and high stability phase and amplitude measurements for high-
speed scanning interferometer applications. It is shown that our new scanning interferometer scheme is very useful
for diagnosing a sample that requires complex analysis. As an example, our new scanning interferometer scheme
has been applied for obtaining phase and amplitude images of the protein biochip samples prepared by using the
sandwich ELISA. The amplitude images are used for diagnosing homogeneity of the sample, while the phase images
are used for measuring the phase difference between samples treated with different concentrations of IL-5. © 2015
Optical Society of America
OCIS codes: (120.3180) Interferometry; (120.5050) Phase measurement; (130.6010) Sensors; (170.3890) Medical
optics instrumentation; (180.5810) Scanning microscopy.
http://dx.doi.org/10.1364/OL.40.002457

Many authors have shown that a two-beam interferom-
eter, either heterodyne or homodyne, can be used for
high-resolution sensor applications [1–8]. In particular,
an in-phase and quadrature (I/Q) interferometer is ideal
for sensor applications because of the following reasons:
(1) it can measure both the phase and amplitude change
induced on the probe beam (PB); and (2) it can measure
the phase difference between the PB and the reference
beam (RB) without any calibration procedure [9,10]. This
scheme is particularly advantageous in such applications
as the scanning microscopy, biosensor, and so forth,
because it can provide more information than the
conventional two-beam interferometer [9–11]. Various
I/Q-interferometer schemes that can be used for these ap-
plications can be found in [9–13]. The major drawback in
these schemes, however, as pointed out by Yoon et al.
[11], is the susceptibility to environmental perturbations:
since the sample and reference mirror are located in two
separate locations, it is highly susceptible to environ-
mental perturbations such as atmospheric turbulence,
temperature change, acoustic noise, and so forth, and
vibrations of the sample result in phase noise.
In principle, lateral scanning of the sample is indepen-
dent of the vibrations along the beam-propagation direc-
tion and should not cause any interference. In practice,
however, pitch and yaw of the scanning stage in conjunc-
tion with a spring-loaded adjustable sample mount are
responsible for the vibration noise during the scanning.
The vibration is not only a major source of the phase
noise but also limits the scanning speed. In all, interfer-
ences caused by environmental perturbations and vibra-
tions of a scanning stage have been major drawbacks of
the conventional scanning interferometer scheme.
Recently, Yoon et al. showed that the interferences
caused by environmental perturbations can be minimized
by employing the balanced-path I/Q-heterodyne interfer-
ometer in which the PB and the RB paths are geometri-
cally balanced and running close to each other so that the
sample and the reference mirror (RM) can be mounted
0146-9592/15/112457-04$15.00/0
on the same stage [11]. They were able to obtain ∼19 dB
rejection of common longitudinal motion by mounting
the sample and the RM on the same stage, where the
common lateral motion was deliberately applied. It
should be mentioned, however, that the PB and RB paths
cannot be truly balanced in a heterodyne interferometer,
because they have different frequencies.
In this Letter, we are presenting a new homodyne
I/Q-interferometer scheme, in which the PB and RB are
exactly balanced. It will be shown that the new scheme
can provide a 30-dB rejection of vibration noise in the
scanning interferometer application. In addition, al-
though a homodyne I/Q-interferometer requires two
optical balanced mixers that have a 90° phase difference
with respect to each other and, thereby, add some com-
plexity in optical arrangement, it has the following addi-
tional advantages over the heterodyne scheme: (1) it
does not require two beams with different frequencies;
(2) can avoid 3-dB signal loss in the RF mixing pro-
cedure; (3) signal processing can be performed in the
baseband; and (4) relative intensity noise of the laser
can be rejected by employing the balanced detection.
This new scanning interferometer scheme has been ap-
plied for obtaining the phase and amplitude images of bi-
ochips prepared in different concentrations of antigen,
interleukin 5 (IL-5). It will be shown that both homo-
geneity and the optical height variation of the sample
can be measured. It will be shown that our new interfer-
ometer scheme can easily identify the optical height
difference between the biochip treated with 1-pg/ml
and 0-pg/ml concentration of IL-5.
A schematic of experimental arrangement is shown in
Fig. 1. Output beam from a stabilized, single-frequency
laser diode-pumped Nd:YAG laser operating at 1319 nm
is split into two paths by using the beam splitter BS1.
The reflected and transmitted beams at the BS1 are used
as the RB and PB, respectively. The plane of polarization
of the output beam from the optical isolator (OI) is
properly oriented at the angle that gives the maximum
© 2015 Optical Society of America
2458 OPTICS LETTERS / Vol. 40, No. 11 / June 1, 2015
Fig. 1. Experimental arrangement. The portion in the dotted
rectangle is the homodyne I/Q demodulator.
transmission of PB and RB at the polarizing beam split-
ters PBS1 and PBS2. The PB is focused onto the sample,
and the reflected beam from the sample is recollimated
and sent back to the PBS1 along the incident path, for
which a concomitant phase and/or amplitude changes
are induced on the PB by the sample. The RB is reflected
at the RM and sent back to the PBS2. Because of the dou-
ble pass in the quarter-wave plate QWP1, the plane of
polarization of the PB and RB are rotated by 90° and
reflected at the PBS1 and 2, respectively. A half-wave
plate, HWP1, is used to rotate the polarization of the
PB by 90° so that the PB is transmitted through the
PBS2. The orthogonally polarized PB and RB are then
combined at the PBS2 and copropagating along the same
path. The phase and amplitude change of the PB are mea-
sured by using the homodyne I/Q-demodulator, which is
specified as the dotted rectangle in Fig. 1. The details
of operation principle for the I/Q-demodulator can be
found in [9] and will not be discussed in detail. The I/Q-
demodulator consists of two balanced mixers, for which
one of them suffers 90° phase shift between the PB and RB
induced by QWP2. The PBS3 and 4 are oriented at 45° to
the plane of polarization of PB and RB so that they can mix
the PB and RB at the corresponding PBSs. The output sig-
nals from the differential amplifier DA1 and DA2 are re-
ferred as vI and vQ , respectively. These output signals
are digitized by using the A/D converters and processed
in the computer to measure the amplitude and phase
change of the PB. The phase difference between the PB
and RB can be determined by the following operation
in the computer:


v
Δϕ
ϕS − ϕR
tan−1 Q ; (1)
vI
where ϕS and ϕR are the phases of the PB and RB, respec-
tively. Therefore, it does not require any calibration to
extract the phase difference Δϕ
ϕS − ϕR , which is deter-
mined by the difference in the effective optical heights
between the sample and RM. The amplitude of PB and
RB can be obtained by
q
RE PB E RB
v2I  v2Q ; (2)
where R is the responsivity of the PDs, and E PB and E RB
are the amplitude of PB and RB, respectively.
In our new scanning interferometer scheme, in order
to make the interferometer immune to vibrations and the
other environmental perturbations, we let the PB and RB
paths be as close as possible to each other. As already
shown in the previous work [11], any acoustic noise
and atmospheric turbulence that have longer character-
istic lengths than the beam separation, ∼1 cm in the
present arrangement, may cause negligible interferences
on I/Q-measurements of the interferometer. In our
present work, it takes less than 5 min for each scanning
measurement. The average RMS phase noise during the
typical measurement interval is obtained by using the
same procedure shown in Ref. 11. The measured average
RMS phase noise is 8.71 × 10−5 rad.
One of the major noise sources is the vibration of the
sample in a conventional scanning interferometer scheme
operating with the reflection geometry where the PB is
focused onto the sample and reflected back into the inter-
ferometer: pitch and roll of the laterally scanning linear
stage can induce vibrations of the sample along the beam
propagation direction, which can appear as the phase
noise in the scanning I/Q-measurements. In our new
scheme, in order to remove interferences caused by vibra-
tions, the sample and the reference mirror are mounted on
the same scanning stage. The vibrations of the scanning
stage are common for both the sample and the reference
mirror, which results in the same induced phase chirp
on both the PB and RB, and, thereby, the interferences
caused by vibrations of the stage can be rejected. The
common vibration rejection ratio was measured by using
the same method used in [11], and the results are shown in
Fig. 2. The red trace (gray trace in black and white printed
version) represents the vibration measurement result
when the vibrating sample and RM were mounted inde-
pendently, while the black trace in the figure represents
the case of both the sample and the RM were mounted on
the same vibrating stage. The results show that a 30-dB
rejection of common mode vibration is ∼11 dB improve-
ment over those for the case of heterodyne version of the
Fig. 2. Frequency spectra representing common vibration
rejection: the gray (red online) and black trace represent the
frequency spectra when the sample mirror is mounted
independently and together with the RM on the PZT stage,
respectively.

Fig. 3. Protein biochip: (a) the schematic of cross-sectional
structure and (b) the SEM image of the bare gold pattern
(the spacing between each tick mark is 10 μm).
interferometer, which has 820-MHz frequency imbalance
between the PB and RB. As a result, we can significantly
improve both the stability and the scanning speed of the
scanning interferometer.
As an example of scanning interferometer applica-
tions, our new interferometer scheme has been applied
for obtaining phase and amplitude images of protein bio-
chips that were prepared on the array of 10 μm × 10 μm
gold patterns by using the sandwich enzyme-linked
immunosorbent assay (ELISA). The samples were fabri-
cated in the Korea Research Institute of Bioscience and
Biotechnology (KRIBB) for other purposes. The samples
used in this work are the bare gold pattern, biochips
treated with 0, 1, 10, 100 pg/ml concentrations of IL-5.
The details of sample preparation will not be given here
because it is out of the scope of this Letter, but the sche-
matic of cross-sectional structure and the SEM image of
the gold pattern are shown in Figs. 3(a) and 3(b), respec-
tively. Each sample and the gold-coated RM were care-
fully aligned and glued to the slide glass that was firmly
mounted on the scanning stage. It should be mentioned
that both the sample and the gold-coated RM can be pre-
pared on an optically flat substrate that can make align-
ment easier and may provide better performance. The PB
was focused and reflected at the gold surface of a sample,
while the RB was reflected at the gold-coated RM. The
spot size of the focused PB was ∼2 μm. The RB was
not focused to the reference mirror to make the scanning
less sensitive to any local defects that may be present on
the surface of the RM. Both the focused PB and the RB
were scanned over a 32 μm × 33 μm area of the sample
surface and RM by use of two axes of a precision motor-
ized 3-axis translation stage (Suruga Seiki D70). It took
less than 5 min to complete the scan.
Figures 4(a) and 4(b) show the phase and amplitude
images of the biochip treated with the 10-pg/ml IL-5 con-
centration. The slope is removed by using proper soft-
ware. No significant defects are observable in these
images. The same scanning measurements were made
on the other samples, for which the images are not shown
here because any significant differences were not observ-
able on these samples except for the optical height
differences between the different samples. The uniform
amplitude images show that the samples are optically
homogenous at the wavelength we used. The line-scan
images, specified as the dotted line in Fig. 4(a), of the
samples with different concentrations are shown in
Fig. 5. The bottom parts and top parts of each curve cor-
respond to the measured phase values at the substrate
June 1, 2015 / Vol. 40, No. 11 / OPTICS LETTERS 2459
Fig. 4. Scanning result showing (a) the phase image and
(b) the amplitude image.
and the pattern. From the bottom to the top (light blue,
blue, green, red, and black curves in color online), the
traces represent the measured phase values of untreated
bare gold pattern, biochips treated with 0, 1, 10, and
100 pg/ml concentrations of IL-5, respectively, which
show clear dependence of measured phases on concen-
trations of IL-5. It should be mentioned that in order to
compare measured phase values at different concentra-
tions, the traces are parallel shifted so that the initial
phase values, which are not the same because of small
but significant height variations introduced in the mount-
ing procedure, are started at the same phase value, 0 rad.
Each trace represents the effective optical height differ-
ence between the corresponding biochip and substrate.
For example, the measured phase difference between the
bare gold pattern and the substrate is 0.93 rad, which is in
good agreement with the theoretically calculated phase
difference, 0.95 rad, for which phase change at the gold
surface and SiN film are taken into account. This result
shows that our proposed scheme can be used as a high-
resolution optical microscope for quantitative analysis
of optical properties in the scanned surface. The details
are not shown here, but dependence of the optical height
on the optical thickness of the sample can be calculated
from the sample structure, and it can be shown that the
phase difference between the substrate and the pattern
increases as the optical thickness of the sample layer
increases. Therefore, it is clear from the results shown
in Fig. 5 that the effective height of a sample treated with
sandwich ELISA is higher than that of the untreated bare
Fig. 5. Line-scan images for untreated bare gold pattern, bio-
chip treated with 0, 1, 10, and 100-pg/ml concentrations of IL-5.
2460 OPTICS LETTERS / Vol. 40, No. 11 / June 1, 2015
Fig. 6. Phase difference between biochip treated with 0, 1, 10,
and 100-pg/ml concentrations of IL-5 and that of 0 pg/ml.
pattern, and the sandwich ELISA pattern treated with a
higher concentration of IL-5 has higher optical height
than that of treated with a lower concentration. The aver-
age phases of the corresponding biochips are obtained by
taking averages of measured phase values of the data
points in the top 80% area of the images, and so it does
for the average phase at the substrate of each sample.
The difference between the average phase of the biochip
pattern and the substrate of each sample is plotted in
Fig. 6. At the present moment, the number of data is
too small to find the exact saturation characteristics in
binding reaction, but the results in Fig. 6 show general
trend in saturation. There may be sudden height change
that may result in π-ambiguity. The amplitude and phase
signals at the boundary between the substrate and sam-
ple, however, show the same dependence as in Ref. 10
which tells us that the average height in the focal spot
varies gradually across the boundary so that the standard
phase unwrapping algorithm can be applied. From the
results shown in Figs. 5 and 6, we can say that our
new scanning interferometer scheme can clearly distin-
guish the difference in optical heights between the
biochip samples treated with 1-pg/ml and 0-pg/ml concen-
tration of IL-5. The average phase difference is 0.07 rad.
It should be mentioned that there are no significant
correlations between the amplitude measurement results
and the IL-5 concentration. The primary reason for this
null result may be the amplitude change due to misalign-
ment. The amplitude change due to a binding reaction at
the wavelength we used is so small that it could be
obscured by the amplitude change caused by a small mis-
alignment in sample mounting. We believe that better am-
plitude measurement results could be available if the
biochips with different concentrations of antigen are
prepared in one substrate. The uniformity in amplitude
image, however, ensures us that the samples are homo-
geneous and have no significant defect.
In summary, a balanced path scheme has been applied
to a homodyne I/Q-interferometer, which exhibits better
environmental noise rejection than that of the heterodyne
interferometer. The interferometer scheme was used for
obtaining the amplitude and phase images of the protein
biochips prepared for various concentrations of IL-5. We
have shown that the true optical height variations of the
biochip samples due to binding reactions can be directly
measured by use of our new interferometer scheme. We
also found that there are strong correlations between op-
tical heights and concentrations of IL-5. We were able to
see clear phase difference, 0.07 rad, between the samples
treated with 1-pg/ml and 0-pg/ml concentration of IL-5,
which is much larger than the average phase noise,
8.71 × 10−5 rad, during the measurement interval. There
are no clear correlations between the amplitude measure-
ment results and the IL-5 concentration. From these
results, we can say that the concentration-dependent am-
plitude change or reflectance change, in other words, at
1319 nm is too small to be distinguished by the amplitude
change caused by misalignments. The uniformity of ampli-
tude image, however, tells us that the samples are homo-
geneously prepared. In all, we have introduced the new
interferometer scheme that can provide both the high-
resolution amplitude and phase images of samples, the
protein biochips in this case. The amplitude images were
used for diagnosing homogeneity of the sample, and the
phase images were used for measuring the changes in
optical heights due to binding reactions. To the best of
our knowledge, it is the first time that both the homo-
geneity and optical height variations of a biochip are diag-
nosed at the same time. We believe that our new scanning
interferometer scheme can be applied for high-resolution
complex analysis of a surface, diagnostics of semiconduc-
tor wafers and devices, high sensitivity readout sensing of
a biochip and fluidic channel, and so forth.
The authors thank Dr. Joohyung Ahn and Dr. Mingon
Kim at KRIBB for providing us the biochip samples. This
research was supported by National R&D Program
through the National Research Foundation of Korea
(NRF) funded by the Ministry of Science, ICT &
Future Planning (NRF-2014M1A7A1A01029956).
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