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Abstract
In the research of resistant aging, the concentration of Growth differentiation factor-11(GDF11) is an indispensable parameter. So the accurate detection of GDF11 is very important in life science and medical cosmetology. Hereby, we proposed and demonstrated a simple method to detect low concentration GDF11 by using fiber surface plasmon resonance (SPR) sensor decorated with two-dimension (2D) material Ti3C2-MXene and gold nanosphere. The sensitivity of the fiber SPR sensor was increased to be 4804.64nm/RIU. After functionalized with GDF11 antibody, the fiber SPR sensor could specifically recognize GDF11, and the limit of detection (LOD) can reach 0.577pg/L which is 100 times lower than that of single-molecule ELISA method.
© 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreement
1. Introduction
Growth differentiation factor-11(GDF11) is an important member of bone morphogenetic proteins (BMPs) subfamily in transforming growth factor - β (TGF - β) superfamily. In 2019, the researchers at Institute Pasteur and CNRS stated the role of GDF11(in blood) in anti-aging [1]. Scientists have reported many research results on the effect of GDF11 [2–6]. They found that GDF11 in blood decreases with age, so the concentration of GDF11 was a very important index. But the existing detection methods need high requirement for instruments, technology and post-processing. So it is necessary to develop a relatively economical and simple sensor to detect GDF11 accurately.
Fiber SPR sensor is widely used in biomedicine [7–9], food safety [10,11], chemical analysis and calibration [12,13] and other fields. Therein, Yang employed fiber SPR sensor to detect DNA Biomolecule [7]. Narsaiah reported a review which included the research results about the application of fiber SPR sensor on food quality and safety assurance [10]. Our laboratory team proposed a fiber microsphere SPR sensor used for the detection of low concentration Hg2+ this year [13]. However, compared with commercial prism-based SPR sensor, the detection sesitivity of fiber SPR sensor should be improved further. A large number of strategies to enhance the sensitivity of the fiber SPR sensor have been proposed and reported. For example, Wang et al. employed the graphene oxide to decorate the sensing silver film of the fiber SPR sensor to increase the sensitivity [14]. And Singh et al. proposed a fiber SPR sensor coated with titanium dioxide for enhanced sensitivity [15].
In recent years, with the development of nanotechnology, nanomaterials are widely used in sensors because of their excellent optical and electrical properties, which can significantly improve the performance of the sensors [16,17]. Initiated by the pioneering work of grapheme, people began to study 2D materials widely for their unique electrical, optical, mechanical, thermal and chemical properties [18]. As a new family member of 2D materials, Ti3C2-MXene not only has excellent adsorption, but also has large specific surface area. In the etching process, the surface of Ti3C2-MXene is easily capped with various functional groups [19,20] without changing its metal conductivity [21]. In addition, Ti3C2-MXene also has obvious SPR characteristics [22], and strong hydrophilicity due to its rich functional groups on the surface. Therefore, Ti3C2-MXene is easy to chemically bond with other ions and macromolecules, which makes it be easily used to connect the sensing film and the substance to be measured. In 2019, Wu and his team innovatively used Ti3C2-MXene to fabricate angle type SPR biosensor to detect carcinoembryonic antigen [23,24]. However, the proposed SPR sensor has a complex structure, and it is difficult to operate. In 2021, Vikas et al. investigated the enhanced performance of the fiber SPR sensor by Ti3C2-MXene, but they only reported the simulating results [25]. In fact, metal nanoparticles can also be used to improve the sensitivity of fiber SPR sensors [26,27].
In this work, by using both Ti3C2-MXene and gold nanosphere together, we designed a novel fiber SPR sensor with higher sensitivity. Furthermore, by decorating the sensor with GDF11 antibody successively, we realize the accurate detection of GDF11.
2. Sensing structure and surface modification
2.1. Fabrication of the sensing probe
According to reference 19], we prepared Ti3C2-MXene by etching Ti3AlC2 in HF acid [19]. The morphology and elements of resultant Ti3C2-MXene were characterized by using scanning electron microscope (SEM) in Fig. 1(a) and energy dispersive spectroscopy (EDS) spectra in Fig. 1(b). From Fig. 1(b) we can find that, the 2D material Ti3C2-MXene is rich in F, O, Al, Cl and other elements. So –O, – OH and –F groups can be formed easily on its surface, and these groups are easy to chemically bond with metal nanoparticles, metal film and the functional groups of the biological protein. Thus Ti3C2-MXene can be used for the connection of the sensing film and substances to be detected. Firstly, we immobilized Ti3C2-MXene on the surface of the fiber to connect the fiber and the sensing gold film which ensured the gold film was sufficiently robust.
By employing a plastic clad multimode fiber (PTIF125/140/ 250HT37, NewPion Photonics) with the core diameter of 125µm, the sensing probe was fabricated. The coating and cladding with the length of 2cm in the middle of the fiber were stripped mechanically to expose the core. The sensing zone was immersed in the piranha solution (H2SO4:H2O2=3:1) for 90 min. Then, we took out the sensing probe and rinsed it several times with distilled water, a clean silica surface covered with hydroxyl groups was obtained. The clean sensing zone was dipped in the Ti3C2-MXene solution (5mg/ml) for 12h at room temperature. The Ti3C2-MXene was immobilized on the surface of the fiber by dehydration condensation reaction. Subsequently, gold film was coated outside the Ti3C2-MXene by employing plasma sputtering instrument [28], and the film thickness was measured by three-dimensional morphology analyzer (NewView7200, Zygo), the structure of the sensor probe is shown in Fig. 2. Figures 2(a) and 2(b) are the radial sections of plastic clad multimode fiber and sensing area, respectively. Figure 2(c) shows the film structure of the sensing zone.
Fig. 2. Structure diagram of the sensing probe, radial section views of (a) plastic clad multimode fiber, (b) sensing zone, (c) film structure of (b).
By using this Ti3C2-MXene-based sensing probe, we carried out the experiment test. The experimental setup is shown in Fig. 3. The light from the source (HL-1000) entered the sensing probe, and the sensing probe was sealed in a chamber. The investigated solutions were injected into the chamber using a programmable microinjection pumper (LSP01-1A, LongerPump), and the waste solutions flowed into the waste reservoir. The output spectrum of the sensor probe was collected by the spectrometer (USB2000+) and sent to the computer for subsequent data processing.
2.2. Optimization of gold film thickness in sensing area
In the Kretschmann structure [29], the evanescent wave decays exponentially in the direction perpendicular to the interface. When we add a 2D material inside the sensing gold film, the sensing performance of the fiber SPR sensor will be influenced, so it is necessary to optimize the thickness of the sensing gold film. The fabrication processes of the sensing probe are shown in Fig. 4(a). For comparative analysis, we also fabricated the pure gold film sensing probe, and its manufacturing processes are shown in in Fig. 4(b).
Fig. 4. Schematic diagram of (a) Ti3C2-MXene-based fiber SPR sensor, (b) pure gold film fiber SPR sensor manufacturing processes.
We fabricated the sensing probes with the gold film thickness of 30nm, 40nm, 50nm and 60nm, respectively. We employed the four sensing probes with different gold film thickness and the pure gold film sensing probe to test the refractive index of the solution, respectively. The refractive index range is 1.333-1.385RIU, the testing results are shown in Fig. 5. In Figs. 5(a)-5(d), with the increase of the refractive index, the resonance wavelength moves to long wavelength direction. In Fig. 5(e), the sensitivity increases first and then reduces with the increase of the gold film thickness, the sensing probe with the gold film thickness of 50nm has the highest sensitivity of 3413.4nm/RIU. Therefore, we selected the 50nm gold film for the following experiments.
Fig. 5. Testing results of the sensing probes with the gold thickness of (a-d) 30nm, 40nm, 50nm, 60nm, (e)relationship between resonance wavelength and refractive index of the solution to be measured.
2.3. Modification of sensing film
In order to bind the GDF11 antibody conveniently on the surface of Ti3C2-MXene by the functional group, we employ Ti3C2-MXene to modify the sensing gold film. The optimized sensing probe was rinsed several times with distilled water, clean sensing zone was dipped in the Ti3C2-MXene solution (5mg/ml) for 12h at room temperature to allow the assembly of Ti3C2-MXene on sensing gold film. The surface of the sensing zone was analyzed by SEM. As shown in Fig. 6(a), the gold film on the fiber surface is smooth. In Fig. 6(b), Ti3C2-MXene can be seen over the gold film. Figure 6(c) is the cross-sectional profile of the sensing zone, and we find the thickness uniformity of the sensing film is appropriate. We employed the modified sensing probe to detect the solution with the refractive index range of 1.333-1.385, the testing results are shown in Fig. 6(d). In which, the resonance wavelength moves to long wavelength with the increase of the refractive index, and the dynamic range becomes wider after the modification of the Ti3C2-MXene.
Fig. 6. SEM images of the coreless fiber surface coated with (a) Ti3C2-MXene + gold film, (b) Ti3C2-MXene + gold film+ Ti3C2-MXene. (c) Cross-sectional profile of (b), (d) SPR testing spectra of (b).
Due to the abundant oxygen or hydroxyl functional groups and complete metal atomic layers of the Ti3C2-MXene, we employed the Ti3C2-MXene to immobilize the gold nanosphere on the sensing zone. As shown in Fig. 7, Ti3C2-MXene was immobilized inside and outside the gold film, the gold nanosphere was immobilized on the sensing zone by Ti3C2-MXene. Because gold nanosphere has the local electric field enhancement effect [27], the sensitivity of the fiber SPR sensor will be increased. In Fig. 8(a), it can be seen clearly that, the gold nanospheres were immobilized on the sensing zone successfully, the diameter of the gold nanosphere is 80nm. As shown in Fig. 8(b), we verified the refractive index sensing performance of this modified fiber SPR sensor.
Fig. 7. Schematic diagram of Ti3C2-MXene + gold film + Ti3C2-MXene + gold nanosphere fiber SPR sensor manufacturing processes.
Fig. 8. (a) SEM images of the sensing zone after the modification of gold nanosphere, (b) SPR testing spectra of (a).
We recorded the resonance wavelength of the testing results and plotted the relationship curve between the resonance wavelength and refractive index of the solution to be measured as shown in Fig. 9. Through comparison and analysis, after the modification of the gold nanospheres, the sensitivity of the sensing probe increases from 4416.79nm/RIU to 4804.64nm/RIU, which demonstrates the sensitivity of the sensor were enhanced by Ti3C2-MXene and gold nanosphere.
Fig. 9. Relationship between resonance wavelength and refractive index of the solution to be measured.
3. GDF11 sensing
Based on the above analysis, we employed the optimized fiber SPR sensor to design a fiber SPR biosensor. The fabrication and detection process of fiber SPR biosensor is shown in Fig. 10. The sensing probe modified by Ti3C2-MXene and gold nanosphere successively was sealed in a chamber. 300 µL (1mg/L) staphylococcal protein A (SPA) was injected into the reaction chamber and maintained for 2 hours at 4 °C. Afterward, the unbound SPA was removed by the Phosphate buffer solution (PBS). The gold nanosphere was decorated by SPA which can form a proper connection between gold nanospheres and antibody, helping the immobilization of the GDF11 antibody on the surface of the sensing zone. 500µL GDF11 antibody (50ug/L) activated by EDC (0.2 mol/L) and NHS (0.05 mol/L) was injected into the reaction chamber and incubated for 4 h at the temperature of 4 °C. GDF11 antibody was immobilized on the surface of the sensing zone. PBS was injected to remove the unbound GDF11 antibody. Then bovine serum albumin (BSA) (10mg/m L) was injected into the reaction chamber and incubated for 10 min to block unspecific active sites on the sensing probe, and the unbound BSA was washed away by PBS. Finally, the fiber SPR biosensor that can detect GDF11 was obtained.
GDF11 antigen (50ug/L) was diluted with PBS, which makes the GDF11 antigen solution with the concentration of 1 ug/L, 100ng/L, 10ng/L, 1ng/L, 100pg/L and 10pg/L is prepared, respectively. Then GDF11 antigen solutions were tested, and the testing curves were shown in Fig. 11. According to Fig. 11(a), the higher the concentration of the GDF11 antigen solution is, the longer the resonance wavelength is, and the lower the resonance light intensity is. As shown in Fig. 11(b), with the increase of the detection time, the resonance wavelength shift increases rapidly at first and then becomes stable gradually. And the lower the concentration of the GDF11 antigen solution is, the larger the resonance wavelength shift is. After processing the data of the detection results above, we presented the relationship between concentration of GDF11 and resonance wavelength by using Fig. 12. In Fig. 12, the abscissa represents the concentration of GDF11 and the ordinate represents the corresponding resonance wavelength. For clarity, we used logarithmic coordinates for abscissa, and the sensitivity can be represented as 8.49 nm/lgC (lgC is the natural logarithm of the GDF11 concentration). We calculated the LOD using the formula: $\textrm{LOD} = \mathrm{\Delta} \lambda /\textrm{S}$ (where $\mathrm{\Delta} \lambda$ is the resolution of the spectrometer, and S is the sensitivity of the sensing probe). It is difficult to calculate the overall average sensitivity due to the large range of detection concentration. So we employed the two low detection concentration to calculate the average detection sensitivity S. And the LOD of the fiber SPR biosensor for GDF11 detection is 0.577pg/L with resolution ($\mathrm{\Delta} \lambda$) of 0.1nm. Compared with single-molecule ELISA method for GDF11 detection [30], the LOD of the proposed method reduced by 100 times.
Fig. 11. (a) Resonance spectra, (b) resonance wavelength shift with the detection time of the Ti3C2-MXene-based fiber SPR biosensor.
Fig. 12. The relationship between the resonance wavelength and the logarithm of the solution concentration.
4. Conclusion
In this paper, we designed a fiber SPR biosensor to detect GDF1. The investigation indicates that, the decoration of sensor probe by Ti3C2-MXene and gold nanospheres can enhance the sensitivity effectively. After the compound modification, the sensitivity of the fiber SPR sensor was increased to 4804.64nm/RIU which was 2.64 times higher than that of the pure gold film sensing probe. After functionalized with GDF11 antibody, the fiber SPR biosensor can detect GDF11 accurately, and the LOD can reach 0.577pg/L which is 100 times lower than that of the other bioassay method.
Funding
Natural Science Foundation of Heilongjiang Province (LH2021A019, F2018027); East University of Heilongjiang Scientific Research Fund (HDFHX210110, HDFHX210111, HDFKYTD202105); Post-graduate innovative research project of Heilongjiang University (YJSCX2021-023HLJU); Fundamental Research Funds for Chongqing Three Gorges University of China (19ZDPY08); Chongqing Key Laboratory of Geological Environment Monitoring and Disaster Early-Warning in Three Gorges Reservoir Area (ZD2020A0102, ZD2020A0103); Science and Technology Project Affiliated to the Education Department of Chongqing Municipality (KJ1710247, KJQN201801217, KJQN201901226); Natural Science Foundation of Chongqing (cstc2018jcyjAX0817, cstc2019jcyj-msxmX0431); National Natural Science Foundation of China (61705025).
Disclosures
The authors declare no conflict of interest.
Data Availability
No data were generated or analyzed in the presented research.
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