Biosensors and Bioelectronics 53 (2014) 459–464

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Sandwich-format electrochemiluminescence assays for tumor marker based on PAMAM dendrimer-L-cysteine-hollow gold nanosphere nanocomposites Ying Zhuo a,n, Guofeng Gui a,b, Yaqin Chai a, Ni Liao a, Kai Xiao a, Ruo Yuan a,n a Key Laboratory on Luminescence and Real-Time Analysis, Ministry of Education, College of Chemistry and Chemical Engineering, Southwest University, Chongqing 400715, China b College of Chemistry and Chemical Engineering, Bijie University, Guizhou 551700, China

art ic l e i nf o

a b s t r a c t

Article history: Received 24 July 2013 Received in revised form 4 October 2013 Accepted 7 October 2013 Available online 23 October 2013

In this work, a novel polyamidoamine (PAMAM) dendrimer-L-cysteine-hollow gold nanospheres nanocomposite was fabricated and used as the promoter for the peroxydisulfate/O2 ECL system to detect the concentration of the tumor marker carcinoembryonic antigen (CEA). Herein, the carboxylterminated PAMAM dendrimers were decorated with L-cysteine (L-Cys) by EDC/NHS coupling chemistry. Then, the hollow gold nanospheres (HGNPs) were employed as effective nano-carriers for the assembly of PAMAM-L-Cys via thiols-Au bonding, which was used for further loading of detection antibody (Ab2) to form the PAMAM-L-Cys-HGNPs-Ab2 bioconjugates. In the presence of target CEA, the sandwiched immuno-structure can be formed between the capture anti-CEA antibodies (Ab1), which self-assembled on deposited gold modified electrode, and the Ab2 on the PAMAM-L-Cys-HGNPs, thereby resulting in a proportional increase in ECL response, due to the significant enhancement of PAMAM-L-Cys-HGNPs toward peroxydisulfate/O2 ECL system. As a result, a sandwich ECL assay for CEA detection was developed with excellent sensitivity of a large concentration variation from 20 fg/mL to 1.0 ng/mL and a detection limit of 6.7 fg mL  1. & 2013 Elsevier B.V. All rights reserved.

Keywords: ECL immunosensor PAMAM dendrimer L-Cysteine CEA Peroxydisulfate

1. Introduction Recently, immunosensors, owing to their advantages of sensitivity, reliability, low-cost for the detection of disease-related biomarker proteins in complex biological matrixes, have attracted much attention from many fields for their promising potential in early disease screening and diagnosis (Yu et al., 2006; Chen et al., 2007; Jie et al., 2011a). Electrochemiluminescence (ECL) is a light emission process in a redox reaction of electrogenerated reactants, which combines the electrochemical and luminescent techniques (Richter, 2004; Bertoncello and Forster, 2009). Possessing the advantages of the selectivity of the biological recognition elements of immunosensor and the sensitivity of the ECL technique, the ECL immunoassays have become one of the most important analytical techniques (Li et al., 2011; Dai et al., 2012; Pei et al., 2013), because of their simple instrumentation, low cost, low background, high sensitivity, wide dynamic concentration response range and portability (Lai et al., 2009; Hu and Xu, 2010). Dai et al. (2012) fabricated an


Corresponding authors. Tel.: þ 86 23 68252277; fax: þ86 23 68253172. E-mail addresses: [email protected] (Y. Zhuo), [email protected] (R. Yuan). 0956-5663/$ - see front matter & 2013 Elsevier B.V. All rights reserved.

advanced ECL immunosensor based on nafion-carbon nanodots nanocomposite films as antibody carriers for the detection of alpha-fetoprotein (AFP). Chai et al. (2012) developed a novel ECL biosensor based on bifunctional aptamer and ABEI-AuNPs for multianalyte assay of small molecule and protein in a biological sample. However, there is still a critical requirement for rapid, sensitive and low-cost detection methods for the early and sensitive immunosensor for cancer biomarkers. Polyamidoamine (PAMAM) dendrimers, the regular tree-like highly branched macromolecules, have multiple branch ends available for further conjugation and unique properties such as a high density of active groups, intense internal porosity, good structural homogeneity and good biocompatibility (Shi et al., 2007; Antoni et al., 2009; Peng et al., 2008; Ma et al., 2007). Above these advantages, PAMAM dendrimers have been widely applied in many fields, such as host–guest chemistry, surface modifications, metal–ion binding, nanoparticle synthesis and so on (Astruc et al., 2010; Frost and Margerum, 2010; Gao et al., 2013). Recently, it has been reported that the introduction of PAMAM dendrimers in ECL reactions would amplify the signal, which has been of interest for the preparation of new sensors. For instance, Lu et al. (2008) reported that the fourth-generation PAMAM dendrimers could enhance ECL of CdS nanocomposite membranes


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by electrochemical deposition method (Jie et al. (2011b) reported that fifth-generation dendrimers used to label with large numbers of CdSe–ZnS quantum dots (QDs) could enhance ECL of QDs and further be used as an amplified ECL signal probe for versatile cell assays. Besides, Ma et al. (2012) reported that PAMAM was employed in RuðbpyÞ23 þ =TPrA system combined with Hg(II)-specific oligonucleotide for the determination of mercury ion. However, to our best knowledge, there is rarely a report about the PAMAM dendrimers in the ECL system of peroxydisulfate/O2. Inspired by the useful applications of PAMAM dendrimers in the fields of biosensor, we reported a highly sensitive and selective ECL immunosensor for the detection of carcino-embryonic antigen (CEA), a popular biomarker associated with liver, colon, breast and colorectal cancer. In our previous works, gold nanoparticles (NPs) and L-cysteine (L-Cys) were used to enhance peroxydisulfate ECL (Wang et al., 2012). Hence, an efficient amplification strategy was proposed by employing hollow gold nanospheres (HGNPs), PAMAM dendrimers and L-Cys as the promoter for the ECL reaction of peroxydisulfate/O2 system. First, L-Cys was conjugated with the PAMAM dendrimers by the formation of an amide link between the carboxyl of PAMAM dendrimers and the amino of L-Cys to obtain PAMAM dendrimers-L-Cys (PAMAM-L-Cys) conjugates. Second, PAMAM-L-Cys were assembled on the HGNPs, which were also used for further immobilization of capture anti-CEA antibodies (Ab2) to form the PAMAM-L-Cys-HGNPs-Ab2 bioconjugates. The results proved that the commendable method enhanced the sensitivity of peroxydisulfate/O2 system. The ECL intensity of the immunosensor increased proportionally with CEA concentration. Thus, this work provided a new method for signal amplification and would extend the application of peroxydisulfate/O2 system in bioanalysis.

2. Experimental 2.1. Reagents and materials Carboxyl-terminated polyamidoamine (PAMAM, G 4.5) dendrimers were obtained from Aldrich (St. Louis, MO, USA). Hydrogen tetrachloroaurate (HAuCl4  4H2O), bovine serum albumin (BSA, 96–99%) were acquired from Sigma-Aldrich (USA). N-(3-dimethylaminopropyl)-N-ethylcarbodiimidehydrochloride (EDC) and N-hydroxy succinimide (NHS) obtained were from Shanghai Medpep Co. (Shanghai, China). Carcinoembryonic antigen (CEA), carcinoembryonic antibody (anti-CEA) γ-glutamyltranspeptidase (γ-GTP) antigen and alpha-L-fucosidase (AFU) antigen were bought from Biocell Company (Zhengzhou, China). K2S2O8 was purchased from Chengdu Chemical Reagent Company (Chengdu, China). L-Cysteine (L-Cys) was purchased from Kangda Amino Acid (Shanghai, China). All other chemicals were of reagent grade and used as received. Phosphate buffered solution (PBS) (pH 7.4, 0.1 M) was prepared with 0.1 M Na2HPO4, 0.1 M KH2PO4 and 0.1 M KCl and used as 3  =4  working buffer solution. Ferricyanide solutions [FeðCNÞ6 ] contained 10 mM K3Fe(CN)6 and 10 mM K4Fe(CN)6 used as a redox reporter, 0.1 M Na2HPO4 and 0.1 M KH2PO4 as ion-paired buffer species (pH 7.4), and 0.1 M KCl as supporting electrolyte. 2.2. Apparatus The ECL emission was monitored by an MPI-A electrochemiluminescence analyzer (Xi'an Remax Electronic Science & Technology Co. Ltd., Xi'an, China) in PBS (0.1 M, pH 7.4) containing 0.1 M K2S2O8 (air saturation) with the voltage of the photomultiplier tube (PMT) at 800 V in detection. Cyclic voltammetry (CV) were conducted with a CHI Instruments Model 660D electrochemical workstation (Shanghai Chenhua Instrument Co., Ltd., PRC). A conventional three-electrode system was used with a glassy carbon electrode

(GCE, ϕ¼4 mm) as the working electrode, a platinum wire as counter electrode and Ag/AgCl (saturated KCl) as the reference electrode in the experiment. The morphology and sizes of various nanomaterials were analyzed using a scanning electron microscopy (SEM) system (SEM, S-4800, Hitachi, Japan) with an acceleration voltage of 10 kV. The value of pH was measured by pH meter (MP 230, Mettler-Toledo, Switzerland). 2.3. Preparation of the PAMAM-L-Cys-HGNPs-Ab2 bioconjugates First of all, hollow gold nanospheres (HGNPs) were prepared according to the literature with a little modification (Schwartzberg et al., 2006). Initially, 400 μL of a solution of Na3C6H5O7  2H2O (0.1 M) was added into 100 mL of double distilled water with rapid magnetic stirring under the nitrogen (N2) atmosphere. Following that, 400 μL of a freshly made NaBH4 (1 M) and 100 μL CoCl2  6H2O (0.5 M) solution were added, respectively. The color of the solution changed from pale pink to gray. Lastly, the 300 μL 0.1 M HAuCl4 solution was slowly added in drops. The color of the solution changed from dark brown to a deep blue. The remaining cobalt nanoparticles left in the solution were oxidized under ambient conditions with rapid stirring for 30 min. Then the HGNPs were collected by centrifugation and washed several times with double distilled water. The resultant HGNPs were dispersed in 5 mL double distilled water. Next, the carboxyl-terminated PAMAM (1.0 mg) were dispersed into 5 mL double distilled water solution by simply sonicating. An appropriate amount of EDC and NHS was added as coupling reagents to activate the –COOH of PAMAM. Then L-Cys 5 mL (0.05 mg/mL) was added in the above solution for combining the carboxyl of PAMAM with the amido of L-Cys to obtain L-Cys functionalized PAMAM (PAMAM-L-Cys). Subsequently, 1.5 mL of the above PAMAM-L-Cys solution was mixed with the obtained 1 mL HGNPs solution to achieve the PAMAM-L-Cys decorated HGNPs composite. Finally, coupling reactions between PAMAM-L-Cys-HGNPs composite and anti-CEA protein were initiated with the aid of EDC and NHS. Briefly, 500 μL Ab2 solution was dropped into 2.5 mL activated PAMAM-L-Cys-HGNPs solution, allowing them to react under softly stirring at 41 C for 12 h. During this process, with the interaction between cysteine or NH3 þ -lysine residues of proteins and the activated –COOH of PAMAM-L-Cys-HGNPs, the Ab2 was loaded onto the nanocomposite to obtain PAMAM-L-CysHGNPs-Ab2 bioconjugates. Following that, the resultant mixture was then purified by dialysis for 1 day at 4 1C using cutoff dialysis tubing with a molecular weight of 8500. The obtained bioconjugates were re-suspended in 2.5 mL of 2 mM PBS containing 1.0 wt% BSA (pH 7.4), and stored at 4 1C for further use. 2.4. Fabrication of the ECL immunosensor First, the bare GCEs were polished with 0.3 and 0.05 mm alumina slurry, respectively, and ultrasonically cleaned with ethanol and double distilled water for 3 min. Then the electrodes were allowed to dry in the air. Following that, electrodeposition was performed in the solution of 1% HAuCl4 to construct the Au nanolayer on the electrode surface with constant potential  0.2 V for 30 s to get deposited Au nanoparticles modified GCE (DpAuNPs/ GCE), which was used for further immobilization of anti-CEA (Ab1). Then the resulting modified electrodes were coated with Ab1 via incubation with 20 μL of Ab1 for 12 h. Unbound antibodies were removed by rinsing with PBS buffer solution. The nonspecific binding sites of the mentioned products were blocked by incubation with 20 μL of 0.25% (w/w) BSA solution. The proposed immunosensor was soaked in 20 μL of a fixed concentration of CEA for 40 min at room temperature. Based on

Y. Zhuo et al. / Biosensors and Bioelectronics 53 (2014) 459–464

sandwich-type immunoreaction, 15 μL of prepared PAMAM-L-CysHGNPs-Ab2 bioconjugates were put on the modified electrodes surface and incubated for 40 min. The resultant electrodes were rinsed with PBS buffer solution at every step. The fabrication procedure of the ECL immunosensor is illustrated in Scheme 1. In the ECL procedure, S2 O28  is reduced to the strong oxidant SOd4  at the surface of the working electrode. Subsequently, partial SOd4  oxidized H2O to form an unstable intermediate HOd which immediately turned into a stable reducing reagent OOHd. OOHd is reduced by SOd4  to produce the excited-state 1(O2)*2, giving the ECL emission (Yao et al., 2008; Wang et al., 2012). The principle of the PAMAM-L-Cys involved in the enhancement of the ECL efficiencies of S2 O28  /O2 occurs as the following: PAMAM-L-Cyse  -PAMAM-L-Cysd þ PAMAM-L-Cysd þ -PAMAM-L-Cysd þ H þ PAMAM-L-Cysd þHOOd-PAMAM-L-Cysþ 1(O2)2* 1

(O2)2*-23O2 þhυ

3. Results and discussion 3.1. Characteristics of the as-prepared HGNPs and Ab2 bioconjugates Scanning electron micrographs (SEMs) were performed to characterize the shape of HGNPs and Ab2 bioconjugates. As we can see, the centers of the HGNPs were dark and the edges were bright (Fig. 1A). It indicated that the morphology of the HGNPs was a hollow structure and the resultant HGNPs were synthesized successfully. Fig. 1B shows a representative SEM image of the Ab2 bioconjugates. The dense coverage of viscous substance on the HGNPs surface could be observed, which supported that PAMAM-LCys-Ab2 has been effectively attached to the surface of the HGNPs.


3.2. ECL and electrochemical behaviors of immunosensor In this work, ECL was used to give detailed information on ECL signal changes of the modified electrodes, shown step by step in Fig. 2A. Compared with bare GCE (curve a), a significant increase in the ECL intensity was observed after deposition in the HAuCl4 solution (curve b), which may be attributed to the excellent electroconductivity of DpAuNPs. Then the ECL intensity decreased successively after assembly of Ab1 (curve c) and blocking reagent of BSA (curve d) because they both hindered the electron-transfer between S2 O28  and the electrode surface. The ECL signal decreased again after the incubation of CEA because the protein also inhibited the electron transfer (curve e). Finally, a remarkable ECL increase was observed after incubation of PAMAM-L-CysHGNPs-Ab2 bioconjugates (curve f). The reason may be that PAMAM-L-Cys-HGNPs could greatly enhance the ECL signal in the ECL system of peroxydisulfate. These results also provided evidence for the existence of PAMAM-L-Cys-HGNPs-Ab2 bioconjugates on the interface. The cyclic voltammogram (CV) of the redox probe of [Fe(CN)6]3 /4 is a useful tool to monitor the barrier of the modified electrode, which is sensitive to surface chemistry, and is employed to indicate the electrochemical behaviors of the sensor at a different stage (Jie et al., 2007). As Fig. 2B shows, a couple of quasi-reversible, well defined redox peaks of [Fe(CN)6]3 /4 were observed on the pretreated bare electrode (curve a). When the gold nanoparticles were first electrodeposited onto the electrode, the redox peak current (curve b) obviously increased due to good conductivity of DpAuNPs monolayer. When the Ab1 was drop-coated onto the DpAuNPs modified electrode, a dramatic decrease in the peak current was noted (curve c). The reason is that the Ab1 can hinder the electron transfer capability of the electrode. After blocking with BSA, the redox peak current was decreased continuatively in turn (curves d). Finally, the modified electrode was incubated with CEA for 40 min, the peak current of curve e decreased further than curve d, because the captured CEA could hinder the electron transfer.

Scheme 1. Schematic diagram of the preparation of the ECL immunosensor.


Y. Zhuo et al. / Biosensors and Bioelectronics 53 (2014) 459–464

Fig. 1. SEM images of HGNPs (A) and Ab2 bioconjugates (B).

Fig. 2. (A) The ECL responses of the modified electrodes step by step: (a) bare electrode, (b) DpAuNPs/GCE, (c) Ab1/DpAuNPs/GCE, (d) BSA/Ab1/DpAuNPs/GCE, (e) CEA/BSA/ Ab1/DpAuNPs/GCE and (f) PAMAM-L-Cys-HGNPs-Ab2/CEA/BSA/Ab1/DpAuNPs/GCE. Scanning from  2.0 to 0 V with a scan rate of 0.1 V/s in 0.1 M K2S2O8 (pH 7.4 PBS). (B) Cyclic voltammograms of the electrode at different stages in 0.1 M PBS (pH 7.4) þ 0.1 M KCl þ2.5 mM [Fe(CN)6]3  /4  solution for (a) bare electrode, (b) DpAuNPs/GCE, (c) Ab1/DpAuNPs/GCE, (d) BSA/Ab1/DpAuNPs/GCE and (e) CEA/BSA/Ab1/ DpAuNPs/GCE. Scan rate of 0.05 V/s and all potentials are given vs. SCE.

3.3. Comparison of the immunosensors using differently labeled Ab2 In order to testify the amplification effect of HGNPs, L-Cys and PAMAM in the S2 O28  /O2 ECL system, we prepared two other sets of biorecognition elements besides the proposed PAMAM-L-Cys-HGNPsAb2 bioconjugates, including HGNPs labeled Ab2 (HGNPs-Ab2) and L-Cys-HGNPs labeled Ab2 (L-Cys-HGNPs-Ab2). All the assay procedures were performed with the same concentration analyte (10 pg/mL CEA) under the same circumstances. As shown in Fig. 3, the immunosensor based on HGNPs-Ab2 bioconjugates (curve b) exhibited higher ECL signal than that of without Ab2 (curve a). The reason may be that the HGNPs in the proposed Ab2 had outstanding electroconductivity, which greatly accelerated the electron transfer between S2 O28  and the electrode surface to obtain more luminescent intermediate SO4 at the unit time. On the other hand, the immunosensor based on L-Cys-HGNPs-Ab2 bioconjugates (curve c) exhibited greater ECL signal than that of curve b; it is because the 2 L-Cys could catalyze the reduction of S2 O8 solution (Niu et al., 2011). However, the proposed PAMAM-L-Cys-HGNPs-Ab2 bioconjugates (curve d) exhibited a much higher ECL signal than that of the others. The reason may be that PAMAM also could improve the ECL efficiencies of S2 O28  and increase the ECL amplifying effect for the modified electrode, resulting in the final enhancement of detection sensitivity. Thus, the proposed Ab2 bioconjugates were adopted for sandwich-type immunosensor to obtain much greater signal amplification in the following experiments. 3.4. ECL response of the immunosensor to CEA concentration The ECL signals relevant to the changes in CEA concentrations are shown in Fig. 4A. The ECL intensity gradually increased with

Fig. 3. The ECL-time profiles of (a) CEA/BSA/Ab1/DpAuNPs/GCE, (b) HGNPs-Ab2/ CEA/BSA/Ab1/DpAuNPs/GCE, (c) L-Cys-HGNPs-Ab2/CEA/BSA/Ab1/DpAuNPs/GCE, (d) PAMAM-L-Cys-HGNPs-Ab2/CEA/BSA/Ab1/DpAuNPs/GCE. Scanning from  2.0 to 0 V with a scan rate of 0.1 V/s in 0.1 M K2S2O8 (pH 7.4 PBS).

increase in the CEA concentrations (Fig. 4A, curves a–g). The ECL signal was proportional to the logarithm concentration of CEA in the large range of 20 fg/mL to 1.0 ng/mL with a regression equation of I (a.u.) ¼ 1928 log cþ 12,341 and a correlation coefficient of 0.994. The detection limit was 6.7 fg/mL and the signal-tonoise ratio was 3. This suggests that this strategy is highly

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Fig. 4. (A) Responses and calibration curve (inset) of immunosensor at different concentrations of CEA. The concentration of CEA (a–g): 20 fg/mL, 50 fg/mL, 100 fg/mL, 500 fg/mL, 1 pg/mL, 10 pg/mL, 1 ng/mL. Scanning from  2.0 to 0 V with a scan rate of 0.1 V/s in 0.1 M K2S2O8 (pH 7.4 PBS). (B) The selectivity of the proposed ECL immunosensor detection of CEA (1.0 pg mL  1) against a different interfering sample: BSA (10 pg/mL), AFU (10 pg/mL), γ-GTP (10 pg/mL) and a mixture (1.0 pg/mL CEAþ 10 pg/mL AFUþ 10 pg/mL γ-GTP), CEA (1.0 pg mL  1). (Error bars: SD, n¼3.) (C) The ECL stability of the immunosensor with the concentration of CEA (10 pg mL  1) under consecutive cyclic potential scans for 14 times.

Table 1 Comparison of the CEA detection with some different ECL emitters. ECL system

Linear range (ng/mL)

Detection limit (ng/mL)



5  10  3–50 3  10  4–10 1  10  3–10

8.2  10–4 6  10–5 8  10–4

Deng et al. (2013) Jie et al. (2010) Zhang et al. (2012)

1  10  4–100

3.3  10–5

He et al. (2013)

1  10–4–40 2  10–5–10

3  10–5 6.7  10–6

Jiang et al. (2013) Present work

RuðbpyÞ23 þ =TPA RuðbpyÞ23 þ =His Luminol/H2O2 S2 O28  /O2

sensitive and has a great potential for early and accurate detection of CEA. Additionally, we also made a comparative study between proposed immunosensor and previously reported immunosensor (Table 1). We can see that the present work for CEA detection based on signal amplification of PAMAM-L-Cys-HGNPs showed a wide linear range response and low detection limit over the compared approach.

was also incubated with 1.0 pg/mL CEA containing 10 pg/mL AFU and γ-GTP, compared with the ECL response obtained from the 1.0 pg/mL CEA only, no significant difference was found. It indicated that the ECL intensity in the presence of CEA was much stronger than that of the others. This indicated that the immunosensor had a good selectivity for CEA. Stability plays an important role in the capability of the immunosensor. Hence, it was examined by employing one immunosensor for consecutive cyclic potential scans for 200 s with the usage of the ECL analyzer. Under continuous cyclic potential scans for 200 s with potential cyclic scanning of 14 times, the relative standard deviation (RSD) of ECL was 1.3%. As shown in Fig. 4c, it indicates that the immunosensor has acceptable reliability and stability. The reproducibility of the immunosensor was evaluated by analysis of the same concentration of CEA (1 pg/mL) using five immunosensors under the same conditions. All immunosensors exhibited closely ECL responses and the RSD of 3.9% was obtained, which meant that the reproducibility of the proposed immunosensor was acceptable. 3.6. Preliminary analysis of real samples

3.5. Selectivity, stability and reproducibility of the immunosensor Possible interfering substances were also tested to evaluate the selectivity of the present immunosensor. Fig. 4B shows the control experiments were performed by using γ-GTP (10 pg/mL  1) and AFU (10 pg/mL) to replace CEA (1.0 pg/mL), respectively. As shown in Fig. 4B, AFU and γ-GTP did not exhibit any obvious increase in signal compared with the BSA (10 pg/mL). The immunosensor

In order to monitor the feasibility of the ECL immunosensor, recovery experiments were performed by standard addition methods in human serum (obtained from the Ninth People's Hospital of Chongqing, China). As shown in Table 2, the recovery (between 100.4% and 103.3%) was acceptable, which suggested that the proposed immunosensor was available for determining CEA in real biological samples.


Y. Zhuo et al. / Biosensors and Bioelectronics 53 (2014) 459–464

Table 2 Determination of AFP added in normal human serum with the proposed immunosensor. Sample number

Add (pg/mL)

Found (pg/mL)

Recovery (%)

1 2 3 4

0.5 5 50 500

0.502 5.16 50.14 506.1

100.4 103.3 100.3 101.2

4. Conclusions In conclusion, we have developed a signal-on ECL immunosensor for the highly sensitive detection of CEA based on S2 O28  /O2 system. This immunosensor achieves an ultralow detection limit of CEA and a wide dynamic range, which can be attributed to the signal amplified strategy of employing PAMAM-L-Cys-HGNPs nanocomposites as the promoter. Besides, HGNPs as effective nanocarriers could provide the active interface for numerous enhancers of PAMAM-L-Cys anchorage and excellent conductivity to promote the electron transfer. Based on the principle of the proposed immunosensor, we expect that a series of ECL immunosensors for other targets could be similarly developed. Acknowledgments This research was supported by the NNSF of China (21275119, 21105081, and 21075100), Research Fund for the Doctoral Program of Higher Education (RFDP) (20110182120010), Ministry of Education of China (Project 708073), Specialized Research Fund for the Doctoral Program of Higher Education (20100182110015) and Natural Science Foundation Project of Chongqing City (CSTC-2010BB4121and CSTC2011BA7003), the Fundamental Research Funds for the Central Universities (XDJK2010C062 and XDJK2012A004), China. References Antoni, P., Hed, Y., Hordberg, A., Nystrom, D., Holst, H., Hult, A., Malkoch, M., 2009. Angew. Chem. 121, 2160–2164.

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Sandwich-format electrochemiluminescence assays for tumor marker based on PAMAM dendrimer-L-cysteine-hollow gold nanosphere nanocomposites.

In this work, a novel polyamidoamine (PAMAM) dendrimer-L-cysteine-hollow gold nanospheres nanocomposite was fabricated and used as the promoter for th...
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