Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439

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Colloids and Surfaces A: Physicochemical and

Engineering Aspects
journal homepage: www.elsevier.com/locate/colsurfa

Detection limit enhancement of fiber optic localized surface plasmon

resonance biosensor by increased scattering efficiency and reduced
background signal
Hyeong-Min Kim , Se-Woong Bae , Jae-Hyoung Park *, Seung-Ki Lee *
Department of Electronics and Electrical Engineering, Dankook University, Yongin 16890, Republic of Korea

H I G H L I G H T S G R A P H I C A L A B S T R A C T

• The effective coverage of AuNPs is

selected on the optical fiber.
• For improvement of scattering effi­
ciency, AuNPs are directly grown on
fiber optic surface.
• Surface etching is executed for reduc­
tion of background in bare surface.
• The enhancement of the LODs is evalu­
ated for several biomarkers.

A R T I C L E I N F O A B S T R A C T

Keywords: To reduce the limit of detection (LOD) in fiber optic localized surface plasmon resonance (FO LSPR) sensors,
Antireflective surface design strategies with high scattering efficiency and low background signal are proposed. Despite the cost
Biosensor effectiveness of fabrication, miniaturization capabilities, and high portability of optic fibers, the LOD is a limiting
Localized surface plasmon resonance
factor for fiber optic sensor applications. To overcome this limitation, we propose methods to increase the
Optical fiber
scattering efficiency and reduce background signal using gold capping and reactive-ion etching (RIE). Specif­
Seed-mediated growth
ically, three strategies are implemented. First, the effective coverage for detection is selected by analyzing the
sensor output according to the density of gold nanoparticles (AuNPs), which have been immobilized on the cross-
section of the optical fiber. Next, the sensitivity is studied by examining changes in scattering efficiency of the
AuNPs grown with different capping times. Third, RIE is performed on the fiber optic surface to reduce the
reflectance in areas where the AuNPs are not fixed, thereby decreasing the background signal. Thyroglobulin,
amyloid beta monomer, and oligomer are measured using the thus-fabricated FO LSPR sensor. The results are
compared with those obtained with a sensor fabricated without using these strategies. The former showed the
lower LOD and higher accuracy for biomarkers of the two.

1. Introduction
The detection of antigens at low concentrations is essential to pre­
vent disease progression with early diagnosis [1]. The demand for
technologies capable of highly sensitive, accurate, and preferably
real-time determination of diseases is rapidly increasing [2]. Over the
past few decades, nanotechnology has been widely applied in biosensing
for detecting low antigen concentrations [3,4], because nanostructures
can overcome the limitations of sensor characteristics with their high
surface-to-volume ratio [5]. For instance, localized surface plasmon
resonance (LSPR) has been representative applicant for highly sensitive,
label-free, and real-time detection of biological species. LSPR is caused

* Corresponding authors.
E-mail addresses: parkjae@dankook.ac.kr (J.-H. Park), skilee@dankook.ac.kr (S.-K. Lee).

https://doi.org/10.1016/j.colsurfa.2021.127439
Received 15 May 2021; Received in revised form 19 August 2021; Accepted 21 August 2021
Available online 26 August 2021
0927-7757/© 2021 Elsevier B.V. All rights reserved.
H.-M. Kim et al.
by the absorption and scattering of light by free electrons that collec­
tively vibrate in noble metal nanoparticles with subwavelength scale of
the light [6]. When the receptor is introduced on the surface of the
nanoparticle, the LSPR principle can be used for immunoassay because
the band by the extinction changes according to variations in the sur­
rounding refractive index [7]. The adsorption of antigens generates a
difference in refractive index. In particular, using optical fibers as sub­
strate for fixation of nanoparticles shows enhancements in terms of
cost-effectiveness, miniaturization, sample consumption, and ease of
handling of biosensors, which has been introduced as the so-called fiber
optic LSPR (FO LSPR) probes [8]. Attempts to achieve the low limit of
detection (LOD) for early detection have recently proceeded in FO LSPR
applications. Martínez-Hernández et al. made multi-layered gold nano­
particle (AuNP) structure by layer-by-layer assembly using poly­
electrolyte [9]. AuNPs and zinc oxide nanoparticles were successively
immobilized on the tapered region of optical fiber [10]. Although these
methods show improved sensitivity, they suffer from limitations asso­
ciated with complexity and reproducibility in regard to fabrication
processes. Also, these techniques are confined to the modification of
metal nanoparticles. Therefore, methods that can simply adjust nano­
particles and further modify other parts except for nanoparticles are
demanded for the development of FO LSPR biosensor which is detect­
able to the small quantity of antigens.
For the accomplishment of a low LOD in biosensor applications, we
easily induce the increase of scattering efficiency in AuNPs by in situ
controlling the particle size on an optical fiber. In addition, surface
texturing is applied on the fiber optic cross-section for the diminishment
of the background signal. Owing to their usefulness in clinical studies
and early interventions, biosensing devices should exhibit LODs of the
order of pg/ml [11]. LSPR-based sensors are essentially refractometers
[12]. The LOD is commonly given by the smallest detectable difference
in refractive index with reliable results [13]. In other words, the sensi­
tivity, which is defined as the rate of change in the output according to
the refractive index, needs to be improved to reduce the LOD [14]. In
addition, the sensitivity is directly related to the LOD as: LOD = 3σ/S
[15], where σ is the standard deviation of the measured values and S is
the sensitivity in the used sensor. It is known that the sensitivity of the
LSPR sensors is closely related to the scattering efficiency of noble metal
nanoparticles [16]. The reduction of background signal is also associ­
ated with enhanced sensitivity because it makes the signal easier to
distinguish [17].
This study focuses on three strategies to enhance the LOD in an FO
LSPR biosensor. First, the density of AuNPs on the cross-section of the
optical fiber was optimized. The sensitivities of FO LSPR sensors with
different nanoparticle coverages were analyzed. Second, the scattering
efficiency of the nanoparticles were improved through a gold-capping
process that increases the size of the AuNPs. This also ensures the
durability of the AuNPs immobilized on the fiber optic surface. Third,
reactive-ion etching (RIE) was conducted on the non-blocked surface by
nanoparticles. AuNPs act as a mask. Because of the decreased reflectance
in the roughened surface owing to the etching process, the background
signal by the reflection in the bare surface reduced. That is, the scat­
tering efficiency of AuNPs on the optical fiber was controlled and
improved, and the background from the bare surface without nano­
particles decreases through three methods. To test the effectiveness of
the applied strategies, the sensitivities were evaluated for each fabri­
cation process. Finally, the proposed FO LSPR sensor was employed to
measure thyroglobulin (Tg), amyloid beta (Aβ) monomer, and oligomer
to determine whether the LOD is enhanced. Gold capping and surface
etching were conducted in preliminary research, but the focus was to
evaluate the fabrication potential [18]. Here, we measured the impact of
strategies on sensitivity, quantitatively detected biomarkers, and eval­
uated enhancements. The clinical applicability of the proposed structure
is assessed here based on the results.
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Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439
2. Materials and methods
2.1. Materials
Sulfuric acid (H2SO4, ≥ 95.0%), hydrogen peroxide (H2O2, ≥
34.5%), sodium citrate dihydrate (C3H4OH(COONa)3⋅2H2O, ≥ 99.0%),
isopropanol (C3H8O, ≥ 99.5%), and glycerol (C3H8O3, ≥ 99.0%) were
purchased from Daejung Chemicals, Republic of Korea. 3-(ethox­
ydimethylsilyl)propylamine (APDMES, C7H19NOSi, 97%), gold (III)
chloride trihydrate (HAuCl4⋅3H2O, ≥ 99.9%), and bovine serum albu­
min (BSA, ≥ 98%) were supplied by Sigma Aldrich, USA. Gold etchant
(TFA) was purchased from Transene, USA. Borate buffer (pH 8.5, 50
Mm) was obtained from Bioworld, USA. The antibody and antigen of Tg
were obtained at a commercial kit of Shinjin Medics, Republic of Korea.
Purified anti-β-amyloid, 1-8 antibody (SIG-39110), Human β-amyloid
peptide (1-42) were gained in BioLegend, USA. The Aβ oligomer was
prepared by incubating the monomer solution in a convection oven of
37 ◦ C for 6 h. The optical fiber (FG105LCA, 105-μm core diameter) was
acquired from Thorlabs, USA. For the preparation of aqueous solutions,
we used deionized (DI) water with a resistivity of 18.2 MΩ cm (Aqua
MAX™-ultra, Youngin Chromass, Republic of Korea).
2.2. Synthesis of AuNPs
The so-called Turkevich method, which chemically synthesizes the
spherical AuNPs by reducing gold ions with citrate ions, was used in this
study [19]. The gold colloidal solution is prepared by adding sodium
citrate dihydrate aqueous solution to a gold chloride trihydrate aqueous
solution. The carboxyl groups in sodium citrate reduce the gold ions in
the gold chloride trihydrate solution, leading to the organization of
small needle-like crystals. The AuNP size gradually increased as citrate
ions adsorbed on the surface the crystals. When growth stops owing to
the consumption of most citrate ions, an inter-particle repulsive force
acts due to the negative surface charge in carboxyl groups, resulting in a
certain inter-particle distance [20].
We set the target diameter of AuNPs at approximately 50 nm. This
condition was confirmed that ensure nanoparticle’s reproducibility and
robustness for fabrication environment in fiber optic configuration
based on the previous study [21]. The synthetic process is as follows. A
total of 50 ml of 0.01% gold chloride trihydrate solution was vigorously
stirred in an oil bath at 105 ◦ C. Then, 0.5 ml of 1% sodium citrate so­
lution was added and then reacted for 20 min. The colloidal solution was
cooled to room temperature with stirring and stored at 4 ◦ C until use.
The ultraviolet–visible spectrum (UV1800, Shimadzu, Japan) of the
prepared colloid is displayed in Fig. S1.
2.3. Fabrication of FO LSPR sensor with improved scattering efficiency
and reduced background signal
The schematic diagram of the fabrication of the FO LSPR sensor with
gold capping and RIE for improved scattering efficiency and reduced
background signal is shown in Fig. 1. The fabrication process largely
involved the immobilization of AuNPs (Fig. 1(a) and (b)), capping of
nanoparticles based on seed-mediated growth (Fig. 1(c) and (d)), and
bare surface etching by RIE (Fig. 1(e)). The attachment technique of
AuNPs on the fiber optic end facet has been reported in previous
research [22]. Briefly, the optical fiber was firstly cut (S326, OFS Fitel,
USA) for a clear cross-section in the step of AuNP fixation. Then, Piranha
treatment (3:1 mixture of sulfuric acid and hydrogen peroxide) was
performed for surface cleaning. To functionalize amine groups for the
adsorption of AuNPs [23], the optical fiber was immersed in a 5%
APDMES solution diluted in isopropanol (Fig. 1(a)). Finally, nano­
particles were attached onto the optical fiber with a colloidal solution
(Fig. 1(b)). The density of particles on the fiber optic surface was
adjusted by varying the treatment times in the APDMES and gold
colloidal solutions. A field-emission scanning electron microscopy
H.-M. Kim et al.
Fig. 1. Schematic diagram of the fabrication of FO LSPR sensor for improved scattering efficiency and reduced background signal. The capping process for the
growth of AuNPs and surface etching by RIE are applied.
(FE-SEM, S5200, Hitachi, Japan) image of AuNPs immobilized on a fiber
optic tip-end is shown in Fig. S2. The average size of nanoparticles was
44.6 ± 5.3 nm, and small aggregation was found on the end face of the
optical fiber.
To improve the scattering efficiency of nanoparticles, seed-mediated
growth-based gold capping was performed [24]. The simplest way of
enhancing the scattering efficiency is to increase the size of the nano­
particles [25]. We grew nanoparticles on the optical fiber using the
already-immobilized particles as seeds. Firstly, seeds were coated by a
reductant (carboxyl groups in sodium citrate dihydrate) to reduce gold
ions on the surface of AuNPs (Fig. 1(c)). Then, the capping process for
bigger nanoparticles was carried out and completed in a gold chloride
trihydrate solution containing the gold ions (Fig. 1(d)). The concentra­
tions of sodium citrate dihydrate and gold chloride trihydrate aqueous
solutions were 0.5 mM and 1 mM, respectively.
To weaken the background signal, RIE (Plasmalab 80 Plus, Oxford
Instruments, Germany) was implemented on the surface of the bare
optical fiber to which AuNPs are not fixed. The capping method is used
to grow particles on an optical fiber causes physical adsorption between
the fiber and nanoparticles, which allows the successful act of the AuNPs
as a mask [26]. The area for the transmission of incident light was also
protected by particles. In the FO LSPR sensor output collected from the
reflective structure in which nanoparticles are formed on the fiber end
face, the background signal reflected from the bare surface as well as the
scattered signal from the AuNPs is always present. If the background
signal is diminished to isolate only the LSPR output, the quantification
ability of the sensor is enhanced [27]. Reflectance is known to decrease
by building a subwavelength structure (antireflective structure) on a
substrate using RIE [28,29]. We executed dry etching (O2/CF4 of
5 SCCM/50 SCCM, power of 150 W, pressure of 55 mTorr) on the fiber
optic surface (Fig. 1(e)).
2.4. Optical system for FO LSPR sensor measurement
Our simple optical system consisted of a light source, a detector, and
a 1 × 2 optical fiber coupler (Thorlabs). One end of the coupler had no
connector and was spliced to the FO LSPR sensor (A-80S, Walfront,
China). In side of two ends, one end was joined to the light source and
the other to the detector. The input from the source excited AuNPs on
the fiber optic surface and scattered signal collected at the detector
through a coupler [30]. When the spectra were observed, white light
(LS-1, Ocean Optics, USA) was used as a source, and a spectrometer
(SM200, Spectral Products, USA) as a detector. When monitoring im­
mune responses, a laser (Iflex-2000, Qioptiq, UK) and a photodetector
(PDA36A, Thorlabs) were used for real-time visualization of the signal.
To minimize external influences, the measurements were conducted
with the fluidic chip in which the FO LSPR sensor was inserted in the
fluidic channel. A schematic diagram of the system is shown in Fig. S3.
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Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439
2.5. Measurement protocol the immune response
The antibody–antigen reaction measurement proceeded within the
fluidic channel. The fluidic channel provides convenience, low sample
consumption, and a stable measurement environment [31]. Before the
antibody–antigen interaction, the antibody and blocking agent binding
treatments were performed on the surface of the particles. The FO LSPR
sensor surface was flushed with DI water for at least 5 min to stabilize
the signal before the antibody binding. Next, the antibody solution
diluted to 20 μg/ml in a borate buffer was injected into the channel. The
supply was continued for 15 min at a flow rate of 10 μl/min. Antibodies
with amine groups are ionically adsorbed on the surface of negatively
charged nanoparticles [32]. Antibodies with amine groups reveal their
positive charge in the buffer solution. Sufficient washing was performed
to remove unbound antibodies. Then, a 1% BSA aqueous solution was
supplied for 15 min to prevent nonspecific binding except for the anti­
body–antigen reaction [33]. The excess BSA was thoroughly removed
for at least 5 min in a DI water atmosphere. Finally, an immune response
was elicited, which ended within 5 min. After the termination of the
immunoassay, the outputs of the sensor were recorded in DI water
before and after antigen binding. The net change of the signal observed
in DI water before and after the immune response is defined as the
output by antigen binding. The output changes recorded in real-time
during a series of reactions for immunoassay are posted in the Fig. S4.
Owing to the miniaturization of the optical fiber, the entire measure­
ment of the immune response takes less than 1 h [34]. The conditions
regarding reaction time and concentration have been adapted from
previous research [35].
3. Results and discussion
3.1. Optimization of nanoparticle density
The signal intensity should be sufficient to distinguish the target
from the background [36]. The dense coverage of AuNPs on the optical
fiber causes an enhanced interaction between the nanoparticles and the
incident light, and this increases the amount of scattered light [37].
Increasing the number of mediators (AuNPs) can also improve sensi­
tivity by the expanded sensing area. We varied the density of immobi­
lized AuNPs on the fiber optic cross-section and analyzed the effect of
the density in the sensitivity before gold capping and surface etching are
introduced. The coverage is adjusted by controlling the amine func­
tionalization time/particle fixation time (Fig. 1(a)/(b)). Both amine
functionalization and particle fixation times can affect final AuNP den­
sity on an optical fiber but the amine functionalization time was
adjusted to be the same as the fixation time of particles for the proper
management of conditions for the FO LSPR fabrication. The amine
functionalization time/particle fixation time was increased from
H.-M. Kim et al.
60 min/60 min to 120 min/120 min with an interval of 15 min. Then,
the samples with 60 min/60 min, 75 min/75 min, 90 min/90 min,
105 min/105 min, and 120 min/120 min were fabricated. Fig. 2(a) il­
lustrates the measured density based on the FE-SEM pictures according
to different fixation times. FE-SEM images of particle distributions on
optical fiber according to different immobilization conditions were
given in Fig. S5. The density progressively increased with increased
fixation times. The changes in the sensor output were measured in
samples with different coverages (Fig. 2(b)), where the refractive index
was increased from 1.33 to 1.38 with an increment of 0.01. The solution
having various refractive indices was prepared by mixing DI water and
glycerol with different ratios. In each sample, sensitivities (slopes) were
displayed in the inset of Fig. 2(b). Sensitivity is defined as the ratio of the
change in LSPR output at resonance peak versus the variation in the
refractive index. Although an upward trend in sensitivity was observed
owing to the increased density of AuNPs, the sample with a density of
31.5% (fixation condition of 105 min/105 min) was the most sensitive
(Fig. 2(c)). This is interpreted to be due to a partial decrease in sensi­
tivity owing to aggregation between nanoparticles in the over-dense
sample (fixation condition of 120 min/120 min) [38]. In the subse­
quent experiments, the amine functionalization time/particle fixation
time was set to 105 min/105 min.
3.2. Modification in capping time of AuNPs
To increase the scattering efficiency and improve the stability of the
nanoparticles on the optical fiber, gold-capping process is conducted.
The enhancement in scattering efficiency was confirmed by measuring
the sensitivity in the samples with different growth times. A reductant
was treated on the optical fiber to which AuNPs were immobilized for
5 min, and the nanoparticles were then grown for up to 120 min in a
solution containing gold ions. Spectra were periodically observed during
growth. In the growth process of AuNPs by capping, the LSPR intensity
was gradually strengthened with larger AuNPs (increase in the
coverage) and improved scattering efficiency. The recorded spectra ac­
cording to the capping time are shown in Fig. S6. Because the increase in
LSPR intensity is insignificant after a capping time of 120 min, it is
believed that the growth is completed with large consumption of the
reducing agent. After the capping process for 120 min, the mean
diameter of the nanoparticles increased to 64.0 ± 6.5 nm with a ratio of
0.16 nm/min. The magnitude of the standard deviation in the particle
diameter was maintained even after the growth procedure [39]. FE-SEM
photographs under different growth times and the correspondingly ob­
tained changes of AuNP density were presented in Fig. S7. Since the
capping process was applied after AuNPs were fixed on the surface of the
optical fiber, an increase in the size of the nanoparticles was observed,
but the rearrangement and the change in distribution were insignificant.
In Fig. 3(a), the sensitivities in three samples with capping times of
Fig. 2. Optimization of fixation time: (a) nanoparticle density at the optical fiber surface according to the amine functionalization time/particle fixation time, (b) the
variation of the measured sensor output at various refractive indices with density, and (c) sensitivity (slope) according to the coverage. The slope is obtained by a
linear regression.
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Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439
0 min, 60 min, and 120 min are compared. It was confirmed that the
LSPR intensity increased by approximately 1.91 times owning to the
improved scattering efficiency after the particles were grown up to
120 min. Based on this, the sensitivities were 26.0, 33.1, and 41.0 in
samples that have capping times of 0 min, 60 min, and 120 min,
respectively. It was demonstrated that the sensitivity was improved in
proportion to the gold-capping time (Fig. 3(b)). The particle growth time
in the capping process was fixed at 120 min because the increase in the
sensor output was saturated after 120 min (Fig. 3(c)). Here, the in­
tensities are considered in the resonance peak. The output intensity can
partly represent the morphology of the AuNPs on the fiber optic surface.
3.3. Selection in the etching time of the fiber optic surface
Upon using the reflective structure with the fiber optic tip-end for
measurement, the detector collects both the LSPR signal, which is
scattered from AuNPs, and the background signal, which is reflected
from the bare fiber optic surface [40]. Because the meaningful infor­
mation for LSPR measurement is the scattered light generated from the
AuNPs, the reflected light from the bare surface (background signal)
should be reduced as much as possible [41]. An antireflective surface is
formed by RIE on the fiber optic end face, where capped AuNPs function
as a mask. The etching time was 5 s, 10 s, and 15 s, respectively. The
etching process is performed only for a maximum of 15 s because
immobilized particles on the surface are lost if it exceeds 15 s. The
surface image of the FO LSPR sensor which was etched for more than
20 s was provided in Fig. S8. The roughness (μSurf, KORTherm Science,
Republic of Korea) of the fiber optic end facet was determined for each
etching time (Fig. S9) [18,42]. The surface roughness was measured
after AuNPs are removed from the surface by an etchant because the
presence of particles can affect the roughness. The background signal
was also recorded in the absence of AuNPs, and it was confirmed that the
background decreased in proportion to the etching time (Fig. 4(a)). The
increase in the etching time makes the surface rougher, which weakens
the reflectivity from the surface where nanoparticles are not present
(background signal) [43]. In Fig. 4(a), the sensor output increased with
the lengthened etching time. At this time, background is gradually
decreased. The sensor output is the LSPR intensity at the resonance peak
observed in the proposed FO LSPR sensor. The background is to record
the signal which is reflected from the bare cross-section of an optical
fiber after surface treatment and AuNP removal. The increased rough­
ness in the fiber optic cross-section reduces the reflection of the signal
from any direction, such as the direction that goes out from the bare
surface and the scattered signal that enters the optical fiber again. In
other words, the reflected background signal from the bare fiber optic
surface without AuNPs is reduced, and the reflection of the scattered
light at which the optical fiber enters from the outside also decreases
(the increase in the LSPR collection from AuNPs), which has the effect of
H.-M. Kim et al. Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439

Fig. 3. AuNP growth time (capping time) in samples with different capping times, (a) variation in the measured sensor’s output with the refractive index and (b)
obtained sensitivities based on the linear fitting, (c) the variation of the intensity with the capping time in one sample.

Fig. 4. Results by RIE on the FO LSPR sensor surface: (a) LSPR output and background signal, which are measured according to the surface etching time, (b) the
variation of the FO LSPR sensor output with the refractive index, and (c) variation in the measured sensitivity with the etching time. The background is monitored
with the removed state of AuNPs after surface etching.

increasing the LSPR intensity to be observed on the detector. The effect
of the decreased background on sensitivity is evaluated in Fig. 4(b) and
(c). The FO LSPR sensor with three strategies responded with the coef­
ficient of determination (R2) of 0.981, which means that high correla­
tion and linearity between sensor output and change in refractive index.
R2 was also obtained based on linear regression with the slope. As a
result of assessing the sensitivities in each sample based on the measured
value in various refractive indices, it can be seen that the sensitivity
improved in proportion to the etching time of the fiber optic surface.
Therefore, it is desirable to select a 15 s etched FO LSPR sensor for the
enhancement of the LOD.
In summary, the condition for immobilization of AuNPs on the op­
tical fiber (density of nanoparticles) for high sensitivity was determined.
In addition, the scattering efficiency was improved through a gold-
capping process that increases the size of the particles, and the re­
flected background from the smooth surface in the optical fiber was
reduced by performing an etching process that roughens the surface. The
improvement in the sensitivity was estimated when each strategy was
applied. Since the efficiency of sensitivity increase by surface treatment
was the most dominant with approximately 2.9 times, the background
relief by the surface etching will mainly contribute to the LOD
improvement. As a result, the immunoassay is proceeded by an FO LSPR
sensor with conditions of the amine functionalization time/particle
fixation time of 105 min/105 min, a capping time of 120 min, and an
etching time of 15 s.
3.4. Confirmation of LOD enhancement
The immune responses are detected in the fabricated FO LSPR sensor
with and without the capping and etching process, respectively. This is
to determine the degree to which the LOD has been enhanced with the
proposed methods. The differences in the measured signal in DI water
before and after the antibody-antigen reaction are shown in Fig. 5. As
targets, Tg, Aβ monomer, and oligomer were chosen. Tg and Aβ are
widely known to be specific biomarkers for thyroid cancer and Alz­
heimer’s disease, respectively. A low LOD is required when monitoring
for recurrence because Tg should not be produced at all in patients who
have undergone thyroidectomy [44], and the presence of even small
amounts of Tg indicates the recurrence of thyroid cancer. The reference
value of Aβ for determining Alzheimer’s disease is several hundred
pg/ml, and a lower LOD is needed for early diagnosis [45]. Each con­
centration is repeatedly detected three times to compensate for the
uncertainty in the measurement. A total of five concentrations of Tg
(from 0.1 pg/ml to 1000 pg/ml) were measured by two types of FO
LSPR sensors. The means and standard deviations of the measured
values by antigen binding are illustrated in Fig. 5(a). It was confirmed
that there were differences in the intensity measured by the FO LSPR
sensor with strategies such as AuNP density optimization, scattering
efficiency improvement, and background reduction being higher than
results obtained by the sensor without the strategies. In the curves of
Fig. 5(a), LODs were obtained on the basis of the linear regression
equation and three times the standard deviations in the measured
values. The LOD was enhanced by a factor of approximately 2.4 times
from 5.1 pg/ml to 2.1 pg/ml when compared to the case before the
introduction of the methods for lower LOD. This trend was also the same
in the measurement results of Aβ monomer and oligomer (Fig. 5(b) and
(c)). The LOD in the monomer became 2.5 times lower, moving from
111.9 pg/ml to 45.1 pg/ml, and the LOD in the oligomer improved by a
factor of 2.2, changing from 33.6 pg/ml to 15.0 pg/ml. These results
indicate the effectiveness of our proposed strategies in the enhancement

5
H.-M. Kim et al. Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439

Fig. 5. Immunoassay results obtained by the proposed sensor and sensor fabricated without gold capping and surface etching: intensity differences gained in DI
water before and after the antibody-antigen interaction in (a) Tg, (b) Aβ monomer, and (c) oligomer, respectively. The various target concentrations were measured.

maximum sensitivity appeared when the time in each step was 105 min.
Table 1
Second, the scattering efficiency and durability were improved by
Comparison of analytical performance for antigens with different biosensor
growing the nanoparticles using the capping process. The LSPR intensity
configurations. MIPs is an abbreviation of molecular imprinting polymers.
and scattering efficiency increased with increase in growth time of
Mechanism Structure Analyte LOD Ref. AuNPs. Particle growth was saturated at a capping time of 120 min.
(pg/ml)
Third, RIE was performed at the optical fiber tip-end using AuNPs as a
Fluorescence Tannylated ferritin Tg 4.3 [50] mask. The surface of the optical fiber was roughened by the etching
nanocage
process, and the reflectivity in the unprotected surface by AuNPs was
Colorimetry MIPs and hemin- Tg 100 [51]
graphene nanosheets lowered accordingly. As a result, the background signal that is reflected
composite in the bare surface without AuNPs was reduced. When the time of the
Electrochemistry Gold electrode on Aβ 140 [52] surface etching exceeded 15 s, AuNPs fell off so the process was carried
nanopillar array monomer out up to 15 s. There were improvements as a result of evaluating the
Ratiometricelectrochemistry Glassy carbon electrode Aβ 500 [53]
monomer
sensitivity of the proposed FO LSPR sensor when each strategy was
LSPR AuNP and etched Tg 2.1 This introduced. The proposed FO LSPR sensor was ultimately applied in
substrate Aβ 45.1 work biosensing applications. Tg, Aβ monomer, and oligomer with various
monomer concentrations were measured to quantify LODs. In the measurements of
the biomaterials, the proposed sensor showed enhanced LODs and CVs
compared to the FO LSPR sensor to which strategies were not intro­
duced. Therefore, by using capping and surface etching, the developed
of LOD. In the proposed sensor, the reduction of the coefficient of FO LSPR sensor with improved scattering efficiency and reduced back­
variation (CV) also contributes to the upgrade of the LOD. The CV is the ground signal can contribute to applications that require a low LOD.
standard deviation/mean as a percentage, which is an index that rep­
resents the spread of the measured values, and it is directly related to the CRediT authorship contribution statement
LOD [46]. After introducing the strategies in the FO LSPR sensor, the CV
decreased by an average of about 8.0%. This is presumed to be due to the Hyeong-Min Kim: Validation, Investigation, Data curation, Writing
slight improvement in the measurement accuracy by the reduced – original draft. Se-Woong Bae: Validation, Investigation, Data cura­
background [47]. In the proposed sensor, a satisfactory correlation of tion. Jae-Hyoung Park: Conceptualization, Methodology, Writing –
the intensity difference and concentration of biomarkers was also ac­ review & editing, Supervision, Resources, Project administration.
quired with an average R2 of 0.930 by linear regression. LODs obtained Seung-Ki Lee: Term, Conceptualization, Methodology, Writing – review
by the proposed FO LSPR sensor are suitable for practical and clinical & editing, Supervision, Resources, Project administration.
applications considering that the reference values of Tg and Aβ are
5 ng/ml (in relapse patient) and 680 pg/ml, respectively [48,49]. Declaration of Competing Interest
In the case of Tg detection, LOD enhanced approximately 2–48 times
compared with other detection methods using fluorescence and color­ The authors declare that they have no known competing financial
imetry in Table 1. For Aβ based on monomer, good performance with interests or personal relationships that could have appeared to influence
respect to electrochemical and ratiometric techniques was as well evi­ the work reported in this paper.
denced considering that recently reported literature only reached the
LOD between 0.14 ng/ml and 0.5 ng/ml. Acknowledgements

4. Conclusion This work was supported by the National Research Foundation of

Korea (NRF) grant funded by the Korea government (MSIT) (Nos. NRF-
In summary, three fabrication conditions were tuned to improve the 2018R1A2B6001361 and NRF-2019R1A2C1083969).
LOD of the FO LSPR sensor. First, the sensor’s sensitivity was optimized
by varying the density of immobilized AuNPs on the fiber optic surface. Appendix A. Supplementary material
To vary the density of AuNPs, the amount of the formed amine group on
the optical fiber was adjusted by the dipping time in the APDMES so­ Supplementary data associated with this article can be found in the
lution, and the amount of electrostatically bound AuNP with amine online version at doi:10.1016/j.colsurfa.2021.127439.
groups was modified by the dipping time in the gold colloidal solution.
As a result of changing the two conditions, it was confirmed that the

6
H.-M. Kim et al. Colloids and Surfaces A: Physicochemical and Engineering Aspects 629 (2021) 127439

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