Letter pubs.acs.org/ac
Cathode Photoelectrochemical Immunosensing Platform Integrating Photocathode with Photoanode Gao-Chao Fan,† Xiao-Mei Shi,† Jian-Rong Zhang,*,†,‡ and Jun-Jie Zhu*,† †
State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, People’s Republic of China ‡ School of Chemistry and Life Science, Nanjing University Jinling College, Nanjing 210089, People’s Republic of China S Supporting Information *
ABSTRACT: Generally, photoanode-based photoelectrochemical immunoassay possesses obvious photocurrent response and lower detection limit for ideal sample detection, but it has the inherent imperfection of poor anti-interference capability for real sample detection. Photocathode-based immunoassay can well avoid the intrinsic drawback of photoanode-based immunoassay, but it has low photocurrent response resulting in less good sensitivity. Herein, a promising new cathode photoelectrochemical immunosensing platform integrating photocathode with photoanode was reported for accurate and sensitive detection of biomarkers. In this proposal, prostate-specific antigen (PSA, Ag) was chosen as a model of target analyte to exhibit the analytical performances of this platform. TiO2/CdS:Mn hybrid structure modified indium−tin oxide (ITO) electrode served as photoanode, whereas CuInS2 microflowers modified ITO electrode was selected as photocathode. The transducer elements of PSA antibody (Ab) were modified on photocathode to fabricate a label-free cathode immunosensing electrode. The proposed immunosensing platform possesses two distinct advantages simultaneously. First, it has good anti-interference capability for the detection of real biological samples, since the biorecognition events occurred on photocathode. Second, the photoelectrochemical system owns evident photocurrent response and low detection limit for target Ag detection thanks to the introduction of the photoanode. Moreover, the proposed immunosensing platform also exhibits good specificity, reproducibility, and stability, and meanwhile it opens up a new horizon to construct other kinds of photoelectrochemical biosensors.
P
includes TiO2, CdS, CdSe, ZnO, CdTe, Bi2S3,6−11 etc., and the p-type semiconductor contains NiO, BiOI, Cu2O, PbS, CuS, and so forth.12−16 Usually, in the photoelectrochemical immunosensing system, the sensing electrode acts as a working electrode, and a Pt wire serves as a counter electrode. Thereinto, anodic photocurrent will be produced if the sensing electrode is constructed on n-type substrate photoactive material (photoande); cathodic photocurrent will be generated if the sensing electrode is established on p-type substrate photoactive material (photocathode).12,14 To date, most of the developed photoelectrochemical immunosensors belong to photoanode-based type.17−22 It is because that photoanode use electrons as the majority carriers, and evident photocurrent response would be produced when electron donor exists, resulting in good detection sensitivity. Besides, n-type semiconductors also own the exciting features of various types, facile synthesis, and easy modification. Yet, photoanode-based photoelectrochemical immunoassay has its inherent imperfection of poor anti-interference capability for
hotoelectrochemical immunosensing is a recently emerged sensor technique, which is developed by the combination of the photoelectrochemical process and electrochemical technology.1 Profitably inherited from the electrochemical method, it has the exciting features of a simple device such as low price, simple operation, and easy micromation, which is very appropriate for fast and real-time biological analysis.2 Moreover, photoelectrochemical detection possesses a high sensitivity due to its low background signal deriving from different energy forms of the excitation source and detection signal.3 Accordingly, photoelectrochemical immunosensing has attracted more and more research interests nowadays. Photoactive materials play a crucial role in analytical performances of the photoelectrochemical immunoassay. Currently, inorganic semiconductor nanomaterials or quantum dots are the most popular photoactive species applied in photoelectrochemical immunosensing field.1,4 According to the types of charge carrier, inorganic semiconductor can be classified into two groups: n-type and p-type.5 Simply put, ntype semiconductor means electrons are the majority carriers and holes are the minority carriers; whereas, p-type semiconductor means holes are the majority carriers and electrons are the minority carriers.5 The n-type semiconductor often used © XXXX American Chemical Society
Received: September 3, 2016 Accepted: October 17, 2016
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DOI: 10.1021/acs.analchem.6b03473 Anal. Chem. XXXX, XXX, XXX−XXX
Letter
Analytical Chemistry the detection of real samples.15,23 As we known, the compositions of real biological samples are very complicated and various reductive agents such as glutathione, dopamine, ascorbic acid, and nicotinamide adenine dinucleotide coexist in them.24−27 When photoanode-based immunosensor is incubated in real samples, the coexisted reductive agents will be inevitably absorbed on the sensing electrode surface, although the sensing electrode is rinsed thoroughly. The absorbed reductive molecules will compete with electron donors in the electrolyte to react with the photogenerated holes at the photoanode/electrolyte interface, leading to unreal photocurrent response of the photoanode.12,15 Owing to intrinsic hole oxidation reaction at the photoanode/electrolyte interface, this trouble is hard to settle by facile change of the ingredient of the photoanode. Photocathode-based photoelectrochemical immunoassay can overcome the inherent drawback of photoanode-based immunoassay. Different from the photoanode, the intrinsic electron reduction reaction occurs at the photocathode/ electrolyte interface, which makes the absorbed reductive molecules from real biological samples have no influence on the reduction reaction, resulting in true photocurrent response of the photocathode itself. And this point has been proved in some previous works such as photocathode-based enzyme sensor and aptasensor.15,23 Unfortunately, the works on photocathode-based immunosensor has not been reported, because photocathode uses holes as the majority carriers and it needs electron acceptor rather than electron donor, resulting in very poor photocurrent response and low detection sensitivity. To play the advantages of both photoanode-based and photocathode-based immunoassay, a new cathode photoelectrochemical immunosensing platform integrating photocathode with photoanode was first proposed in this letter, and the experimental details were described in the Supporting Information. Herein, prostate-specific antigen (PSA, Ag) was selected as an example of target analyte to state practicability of the platform. As illustrated in Scheme 1, a label-free
ITO electrode, which served as photoanode of the photoelectrochemical system. For the cathode immunosensing electrode, CuInS2 microflowers synthesized by the solvothermal method were first coated on a clean ITO electrode, and the dense film was formed after being treated with high temperature. Then, PSA capture antibodies (Ab) were modified onto the ITO/CuInS2 photoanode via the assistance of chitosan linking molecules. After bovine serum albumin (BSA) blocked unbound active sites, the sensing electrode was ready and target Ag was quantitatively determined by evident photocurrent change caused by specific immunoreaction between target Ag and its Ab.
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RESULTS AND DISCUSSION Figure 1A shows powder X-ray diffraction (XRD) pattern of the synthesized CuInS2 products, which is in accordance with monoclinic CuInS2 with the cell parameters of a = b = 5.523 Å, and c = 11.141 Å (JCPDS 27-0159). The observed peaks were assigned to diffractions from the (112), (004), (204/220), (312), and (332) crystal faces and no characteristic peaks from other impurities such as In2S3 and Cu2S were detected, indicating successful formation of pure crystalline CuInS2. Figure 1B exhibits a typical scanning electron microscopy (SEM) image of the CuInS2 products. It could be seen that the samples were microflowers with size distribution from 1.5 to 2.0 μm, and the surface morphologies of these microflowers were composed of nanosheets with about 30 nm in thickness. The rough surface of the products offered more interface area for photogenerated electron exchange, which could effectively increase the photocurrent intensity. For photoelectrochemical immunoassay, enhanced photocurrent response is preferable, since it is directly related to the detection sensitivity. The exploration on optimal fabrication parameters of the photoanode (ITO/TiO2/CdS:Mn electrode) and photocathode (ITO/CuInS2 electrode) was described in the Supporting Information. To illustrate excellent photoelectrochemical property of the system involved in both photoanode and photocathode, control experiment was carried out, as shown in Figure 2A. Curve a presents photocurrent of the photocathode-based system (in which the ITO/CuInS2 electrode acted as the working electrode, and a Pt wire served as the counter electrode), and the response value is very small (curve a, I = −0.28 μA). Curve b displays photocurrent of the photocathode-photoanode-based photoelectrochemical system (in which the ITO/CuInS2 electrode acted as the working electrode, and the ITO/TiO2/CdS:Mn electrode served as the counter electrode), and the response value dramatically enhanced (curve b, I = −227.12 μA), which is close to 3 orders of magnitude better than that of photocathode-based system. The photocurrent of the photoanode-based system was also tested (in which the ITO/TiO2/CdS:Mn electrode acted as the working electrode, and a Pt wire served as the counter electrode), and the response value is evident (curve c, I = 168.65 μA). However, the photocurrent response value of the photoanode-based system is only about 74% of that to photocathode-photoanode-based system. All these results proved that the photocathode-photoanode-based system possessed outstanding photoelectrochemical property compared to photocathode-based as well as photoanode-based system. Besides, the stability of the photocathode-photoanodebased photoelectrochemical system was evaluated. It can been seen from Figure 2B that the photocurrent intensity nearly remained the same after undergoing more than 20 cycles of
Scheme 1. Proposed Cathode Photoelectrochemical Immunosensing Platform Integrating Photocathode with Photoanode
immunosenor was established on the photocathode and was used as the working electrode; whereas, a photoanode instead of Pt wire was used as the counter electrode. Specifically, TiO2 nanoparticles were first covered on a bare ITO electrode, and then Mn2+ doped CdS (CdS:Mn) nanocrystals were deposited on the TiO2 film by successive ionic layer adsorption and reaction to produce TiO2/CdS:Mn hybrid structure modified B
DOI: 10.1021/acs.analchem.6b03473 Anal. Chem. XXXX, XXX, XXX−XXX
Letter
Analytical Chemistry
Figure 1. (A) XRD pattern and (B) SEM image of the synthesized CuInS2 microflowers.
Figure 2. (A) Photocurrent responses of the photoelectrochemical systems: (a) photocathode-based (PC-based), (b) photocathode-photoanodebased (PC-PA based), and (c) photoanode-based (PA-based); (B) time-dependent photocurrent response of the photocathode-photoanode-based photoelectrochemical system.
produces new midgap centers, which can effectively inhibit electron−hole recombination.32 Besides, CdS has a higher conduction band than that of TiO2, which is in flavor of the injection of photogenerated electrons from CdS to TiO2. Thus, combining CdS:Mn with TiO2 to form TiO2/CdS:Mn hybrid structure could effectively increase the harvest of light energy and depress charge annihilation. When electron donor of ascorbic acid (AA) was offered, the photogenerated holes in the valence band (VB) of the photoanode were neutralized, and accordingly an obvious photocurrent response was generated (shown as curve c in Figure 2A). CuInS2 microflowers modified ITO electrode was used as a photocathode. The photogenerated electrons in the conduction band (CB) of the photocathode were trapped by O2 dissolved in AA electrolyte. CuInS2 is a p-type semiconductor, and it has a narrow band gap value of 1.5 eV, allowing it to utilize the light energy efficiently.33 However, the p-type CuInS2 uses holes as the majority carriers and it needs no electron donor to offer electrons, resulting in a very weak photocurrent response (shown as curve a in Figure 2A). While the photoanode and photocathode coexisted in the system, the photocurrent response dramatically increased (shown as curve b in Figure 2A). Also, the reason can be explained from the following two different aspects. For the photoanode of ITO/TiO2/CdS:Mn, the photocathode of ITO/CuInS2 could supply plenty of hole carriers, which significantly increased the attraction of electron carriers coming from the photoanode, resulting in ultrafast electron transfer and effective inhibition of charge recombination in the photoelectrochemical system.34,35 For the photocathode of ITO/CuInS2, the photoanode of ITO/TiO2/ CdS:Mn could effectively utilize electron donors in the
repeated light irradiation, indicating the designed photoelectrochemical system possessed good stability and was appropriate for establishing an immunosensing platform. The photocurrent-enhanced mechanism of the photoelectrochemical system involved in both photocathode and photoanode is illustrated in Scheme 2. TiO2/CdS:Mn hybrid structure modified ITO electrode served as a photoanode. TiO2 is an n-type, wide band gap semiconductor (∼3.2 eV), which can just absorb the ultraviolet light (
