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Turn-On Detection of a Cancer Marker Based on Near-Infrared Luminescence Energy Transfer from NaYF4:Yb,Tm/NaGdF4 Core-Shell Upconverting Nanoparticles to Gold Nanorods Hongqi Chen, Yingying Guan, Shaozhen Wang, Yuan Ji, Mengqi Gong, and Lun Wang Langmuir, Just Accepted Manuscript • DOI: 10.1021/la502753e • Publication Date (Web): 08 Oct 2014 Downloaded from http://pubs.acs.org on October 10, 2014

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Turn-On Detection of a Cancer Marker Based on Near-Infrared Luminescence Energy Transfer from NaYF4:Yb,Tm/NaGdF4 Core−Shell Upconverting Nanoparticles to Gold Nanorods Hongqi Chen*, Yingying Guan, Shaozhen Wang, Yuan Ji, Mengqi Gong, Lun Wang* Anhui Key Laboratory of Chemo-Biosensing, Key Laboratory of Functional Molecular Solids, Ministry of Education, College of Chemistry and Materials Science, Anhui Normal University, Wuhu 241000, PR China

*Correspondence: Phone: +86-553-3869303 Fax: +86-553-3869303 Email: [email protected] (H. Chen); [email protected] (L. Wang)

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ABSTRACT: A homogeneous immunoassay for the sensitive and selective determination of trace amounts of alpha-fetoprotein (AFP, a cancer marker) by detection in the near-infrared (NIR) region based on luminescence energy transfer (LET) from NaYF4:Yb,Tm/NaGdF4 core−shell upconverting nanoparticles

to

gold

nanorods

(GNRs)

is

presented.

The

carboxyl-functionalized

NaYF4:Yb,Tm/NaGdF4 core−shell upconverting nanoparticles (UCNPs) were excited by a 980 nm continuous wavelength laser, and its emission peak appeared at a near-infrared wavelength (~804 nm). The carboxyl-functionalized upconverting nanoparticles were conjugated with the anti-AFP (Ab1) and acted as donor. GNRs with a high absorption band around 790 nm, which was overlapped the UCNPs emission, were synthesized and acted as the acceptor. The donor (negatively charged) interacted with the acceptor (positively charged) via electrostatic interactions to bring them into close proximity. LET could occur, producing a quenching phenomenon. When the AFP antigens were added into the system, the binding affinity between AFP and Ab1 was stronger than the electrostatic interactions, which released the energy acceptors from the energy donors, interrupting luminescence energy transfer, and therefore, the luminescence was recovered. Based on the restored luminescence, a turn-on optical immunosening system was developed. Under the optimal conditions, the linear range of detection was from 0.18 to 11.44 ng/mL for AFP (R = 0.99), with a detection limit as low as 0.16 ng/mL. The proposed method has also been used to monitor AFP in human serum samples. Therefore, further study based on the NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles-GNRs construction may open the way for a new class of NIR-LET biosensors with wide applications.


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1. INTRODUCTION Over the past decade, rare-earth-doped upconversion nanoparticles (UCNPs) have become more prominent in the biological sciences because of their outstanding luminescent properties which exceed those of conventional luminescent materials.1, 2, 3, 4 UCNPs are characterized by a high signal-to-noise ratio because of a very low autofluorescence background,5, 6, 7, 8 narrow emission peaks, large Stokes shifts, low toxicity, and high chemical stability.2, 9, 10, 11 They can be excited by low power continuous wavelength lasers as the excitation source and even by non-coherent light sources. Furthermore, UCNPs can convert low-energy near-infrared excitation light into an emission at visible or near-infrared (NIR) wavelengths,12, 13 so their emission is less harmful to biological samples and displays greater sample penetration depth than conventional ultraviolet excitation.14 Now the bioassays conducted in the near-infrared light range for applications have attracted special attention of science researcher such as noninvasive and deep penetration of NIR radiation,15 immunoassay,16 bioimaging,17, targeted tumor imaging.22,

23, 24

18

chemical sensing19,

20, 21

and in vitro and vivo

To achieve high photoluminescence efficiency, core-shell

structured nanomaterials25 have been prepared, such as silica/NaYF4:Yb,Er nanospheres. Even though the preparation of NIR-to-NIR core−shell upconverting nanoparticles with high photoluminescence efficiency26,

27

is still an emerging field, these nanomaterials have great

potential in NIR range bioassays.28 Recently, our group has developed an efficient way for constructing sandwich-type turn-off NIR luminescence energy transfer (LET) systems by utilizing NaYF4:Yb,Tm upconverting nanoparticles as the donor while gold nanorods (GNRs) serve as the acceptor. These materials have been used for the detection of Hg2+ and thrombin in biological fluids.29, 30 We questioned whether it would be possible to achieve a convenient turn-on luminescent sensing system.


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Herein, we have prepared carboxyl-functionalized NIR-to-NIR NaYF4:Yb,Tm/NaGdF4 core−shell upconverting nanoparticles with high photoluminescence efficiency that acted as the donor and with alpha-fetoprotein (AFP) antigen serving as the model protein. AFP is an embryospecific glycoprotein associated with the normal growth of the mammalian fetus. It is the most reliable and widely used tumor marker for the diagnosis of hepatocellular carcinoma and yolk sac tumor.31, 32 In this work, we describe the development of a turn-on near-infrared LET system based on homogeneous immunoassay and electrostatic interactions33 for the sensitive and selective determination of trace amounts of alpha-fetoprotein in human serum samples.

2. EXPERIMENTAL SECTION 2.1. Materials and Reagents Poly(acrylic acid) (PAA), 2-(N-morpholino) ethanesulfonic acid (MES), and diethylene glycol (DEG) (99+%) and cetyltrimethyl ammonium bromide (CTAB) were purchased from SigmaAldrich. YbCl3·6H2O was purchased from Jinan Henghua Science and Technology Co., Ltd. (China). N-Hydroxysuccinimide (NHS), Gd(NO3)3·6H2O, NaBH4, AgNO3, Y(NO3)3·6H2O, Tm(NO3)3·6H2O and HAuCl4·3H2O were purchased from Aladdin Reagent Corporation (Shanghai, China). 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC⋅ HCl) was provided by Shanghai Medpep Co. Ltd. Ascorbic acid was acquired from Sangon Biotechnology Inc. (Shanghai, China). Concentrated hydrochloric acid, toluene, ethanol, sodium citrate, and NaF were obtained from Sinopharm Chemical Reagent Co. (China). All of the reagents were analytical grade and used without any further purification. Solutions were prepared


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by appropriate dilution of the stock solution with ultrapure water. AFP antigen and Ab1 were purchased from North Biotechnology Research Institute (Beijing, China). 2.2.Instruments and Characterizations Transmission electron microscopy (TEM, JEOL 2010, operating voltage of 200 KV) was applied to investigate the size and morphology of materials, and scanning electron microscopy (SEM, Hitachi S-4800, operating voltage of 10 kV) was used to examine the elements of the substance. LET spectra were obtained with a Hitachi F-4600 fluorescence spectrophotometer with an external 980 nm laser (Beijing Hi-Tech Optoelectronic Co., China) as the excitation source instead of the xenon source. The UV–visible absorption spectra were obtained with a Hitachi UV–4100 spectrophotometer. The zeta potentials were recorded using a Malvern Zetasizer nano

ZS90

apparatus

(Malvern

Instruments,

Malvern,

United

Kingdom).

Concentrations of Gd and Y were determined on an X Series 2-ICP-MS (Thermo fisher Scientific, USA). 2.3. Synthesis and Modification of NaYF4:Yb,Tm Upconversion Nanoparticles NaYF4:Yb,Tm upconversion nanoparticles were synthesized according to a reported method,34 with some alterations in the procedure. Ultrapure water (2.20 mL), Y(NO3)3 (0.20 M, 2.20 mL), YbCl3 (0.10 M, 0.50 mL), Tm(NO3)3 (0.10 M, 100.00 µL), sodium citrate (0.10 M, 1.75 mL), CTAB (0.10 g) and ethanol (15.00 mL) were gradually mixed together under vigorously stirring. Then, NaF (1.00 M, 6.00 mL) was added dropwise and the solution was stirred for 2 h. To obtain the complex precursor, concentrated nitric acid (1.00 mL) was added. Subsequently, the solution was transferred into a 50 mL autoclave flask and heated at 180 °C for 4 h. The solution was allowed to cool to room temperature naturally. The product was separated by centrifugation and was washed several times with ultrapure water and absolute ethanol.


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The as-prepared NaYF4:Yb,Tm UCNPs were modified with PAA by a ligand exchange method.35 PAA (0.20 g) and DEG (16.00 mL) were heated to 110 °C in a four-neck flask. Then, UCNPs (25.00 mg) were dispersed in toluene and the suspension was injected into the solution. The system was heated to 150 °C for 1.5 h. Hydrochloric acid (0.10 M) was added until the solution was cooled down to room temperature naturally. The resultant mixture was centrifugally separated and washed for several times with ultrapure water. The process of this experiment was completed under vigorously stirring in a nitrogen atmosphere. 2.4. Preparation of NaYF4:Yb,Tm/NaGdF4 Core−Shell Nanoparticles The NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles were synthesized by an improved procedure from the literature.36 The PAA-functionalized NaYF4:Yb,Tm UCNPs (5.00 mg) were homogeneously dispersed in ultrapure water (2.00 mL). Appropriate amounts of Gd(NO3)3·6H2O were completely dissolved in 1.00 mL of ultrapure water and then they were mixed with the UCNPs in a centrifuge tube. The mixture was heated to 60 °C with stirring. After 1 h, the mixture was cooled naturally to room temperature. The product was separated by centrifuging and purified by several washes with ultrapure water. 2.5. Bioconjugation of NaYF4:Yb,Tm/NaGdF4 Core−Shell Nanoparticles. The procedure for the preparation of antibodies conjugated NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles was adapted from literature methodology.37,

38, 39

The carboxyl modified

NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (6.40 mg) were completely dissolved in MES buffer solution(10.00 mM, pH 6.4). EDC and sulfo-NHS were added and this solution was incubated with shaking for 2 h at 30 °C. The activated products were obtained by centrifugation and washed with ultrapure water. Subsequently, the activated NaYF4:Yb,Tm/NaGdF4 core−shell


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nanoparticles were dissolved in PBS buffer solution (2.00 mL, 10.00 mM ， pH 7.4) and appropriate amount of Ab1 were added. The solution was incubated for 24 h at 30 °C. To block any remaining active surface of the donor,40, 41 BSA (200.00 µL, 3.00 mg/mL) solution was added. The solution was incubated with suitable shaking for 0.5 h at 30 °C. The final product linked with the antibody was collected and washed with ultrapure water to remove excess materials. 2.6. Synthesis of GNRs GNRs were synthesized by a modified procedure from the literature.42 To prepare the seed solution, CTAB solution (0.20 M, 5.00 mL) and HAuCl4 (5.00×10-4 M, 5.00 mL) solution were successively added into a flask. Then, ice-cold NaBH4 solution (1.00×10-2 M, 0.60 mL) was gradually added into the mixture under suitable stirring. After 2 min, the seed solution was placed at 27-30 °C. The growth solution was prepared with CTAB (0.20 M, 5.00 mL), AgNO3 (4.00×10-3 M, 0.25 mL), HAuCl4 (1.00×10-3 M, 5.00 mL) and ascorbic acid (7.88×10-2 M, 70.00 µL), which was gradually added into the flask with stirring, resulting in the colorless growth solution. The growth solution was stored at 27-30 °C. An aliquot (12.00 µL) of the seed solution was added into the growth solution. After approximately 40 min, GNRs were collected by centrifugation and washed several times with ultrapure water. 2.7. Luminescence Energy Transfer Detection of AFP The above bioconjugation of carboxyl-functionalized NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (0.80 mg/mL) and GNRs (1.95×10-10 M) were mixed in PBS buffer solution (10.00 mM, pH 7.4). Then various amounts of AFP or other test samples were added into the complex matrixes. Finally, the sample solutions were incubated with shaking for 1.5 h at 30 °C. LET


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spectra were acquired with a Hitachi F-4600 fluorescence spectrophotometer with a 980 nm laser (5.0 nm for slit width and 700 V for PMT voltage).

3. RESULTS AND DISCUSSION 3.1. Characteristics of the NaYF4:Yb,Tm/NaGdF4 Core−Shell Nanoparticles and GNRs During the synthetic process of the spheres of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles, the quantity of Gd(NO3)3·6H2O was optimized to enhance optical properties. At the Gd(NO3)3·6H2O concentration of 5.56×10-2 M, the luminescence intensity reached the maximum (shown in the Supporting Information, Figure S1); ICP-MS data showed that the molar ratio of Gd/Y was 24.47%. The result of ICP-MS data indicated the existence of Gd3+. The TEM, EDS images (Figure 1) and XRD (Figure S2) confirmed that the NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles with properly uniform size were prepared successfully. Figure 2 displays the TEM image of the GNRs. The XRD pattern of the gold nanorods were measured (Figure S3). The emission spectrum of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (a) and the absorbance spectrum of GNRs (b) exhibited large overlaps (Figure 3). We observed sharp and narrow bands in the emission spectrum of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles. This result can be primarily ascribed to electric and magnetic dipole optical transitions with the 4fn energy manifolds and may also implicate 4fn–15d,43 whose positions are weakly connected with the external environment. At the same time, the absorbance spectrum of the GNRs showed two peaks at 530 and 790 nm, which were able to overlap the emission band of the UCNPs. The transverse SPR band of GNRs is a constant value, but the longitudinal SPR band is known to be


8

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located in the near-infrared region and the specific location depends on the morphology of GNRs.44

Figure 1. The TEM image (A) and EDS (B) of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles.

Figure 2. The TEM image of GNRs


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Figure 3. The emission spectrum of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (a) and the absorbance spectrum of GNRs (b).

The Principle of Determining AFP Levels The schematic depiction for AFP determination is shown in Figure 4. PAA is highly biocompatible and readily allows the incorporation of a functional end group. Thus, the NaYF4:Yb,Tm nanoparticles were modified with PAA based on ligand exchange to improve their biocompatibility.45 The zeta potential values varied from 30.60 mV to -22.70 mV after modification with PAA. To enhance their luminescence intensity, the NaYF4:Yb,Tm/NaGdF4 core−shell

nanoparticles

were

prepared.

The

resultant

carboxyl-functionalized

NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles could be easily connected to Ab1, and the zeta potential was found to be -14.50 mV. The above zeta potentials were all measured in PBS buffer


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solution (10.00 mM, pH 7.4). The carboxyl-functionalized upconverting nanoparticles were conjugated with the Ab1 and acted as the donor. GNRs served as the acceptor. The donor (negatively charged) interacts with the acceptor (positively charged, 19.16 mV) via electrostatic interactions to bring them into close proximity. So, LET will occur, producing a quenching phenomenon. When the AFP was added into the system, the affinity between AFP and Ab1 would be stronger than the electrostatic interactions, which releases the energy acceptor from the energy donors. Thereby, the luminescence would be restored.

Figure 4. The schematic depiction for determining AFP.


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Control Experiments To confirm that the specific recognition between the AFP and the Ab1 blocked LET, we conducted a series of control experiments under the optimum conditions (Figure 5.). Ab1modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles exhibited a strong fluorescent signal (a). In the absence of GNRs, when AFP was added (b), we observed no obvious changes in the luminescence intensity, indicating that AFP has no effect on this system. When GNRs were added (c), the donor interacted with the acceptor via electrostatic interactions, bringing the GNRs into close proximity, resulting in high luminescence quenching efficiency. However, when the AFP was added into the solution of Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles and GNRs (d, e), the luminescence intensity of the antibody-modified nanoparticles increased, suggesting that the binding between AFP and its Ab1 was stronger than the electrostatic attraction between the Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles and the GNRs. Therefore, the distance between the donor and the acceptor was increased, which prevented the quenching effect of LET, and fluorescence intensity increased. To ensure that the control experiments sufficient, we studied the effect of ionic strength (concentration

of NaCl) on

the luminescence intensity recovery of Ab1-modified

NaYF4:Yb,Tm/NaGdF4 after quenching with GNRs (Figure S4). The results showed that high ionic strength led to a significant luminescence intensity recovery.46


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Figure 5. Luminescence emission spectra of (a) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles; (b) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + 7.62 ng/mL AFP; (c) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs; (d) Ab1modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs + 2.86 ng/mL AFP; (e). Ab1modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs + 7.62 ng/mL AFP. Conditions: Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles 0.80 mg/mL; GNRs: 1.95×10-10 M; experiments were performed in PBS buffer (10.00 mM, pH 7.4).

Optimization of the Assay We examined a number of factors that could affect the assay results in an effort to improve the sensitivity of this procedure. We investigated the effects of the concentration of Ab1 used during


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the preparation of antibody-modified nanoparticles and determined that 6.25 µg/mL of Ab1 provided the most effective Ab1–modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (Figure 6 A). We examined the effect of the ratio of acceptor (UCNPs) to donor (GNRs) on the detection sensitivity and the linear range of the LET system (Figure 6 B). If the proportion of UCNPs was too high, the sensitivity would decrease, while too great a proportion of GNRs would result in a greater LET and the linear range would become too narrow. These studies showed that the luminescence quenching efficiency reached ~70% at a GNRs concentration of 1.95×10-10 M, so this quantity was chosen in this experiment. Figure 6 C shows that the response of AFP reached the maximum when the concentration of Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles was 0.80 mg/mL, so this concentration of the Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles was used in our work. We also determined that luminescence recovery reached the maximum value with 1.5 h incubation at 30 °C, and remained relatively stable for approximately 30 min (Figure 6 D). Under these conditions, a good recovery rate was obtained.


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Figure 6. (A) Luminescence intensity: NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (0.80 mg/mL) + various concentrations of Ab1. (B) Luminescence quenching efficiency: Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (0.80 mg/mL) + various concentration of GNRs. (C) The influence of different concentration of Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles on the response of AFP with 1.95×10-10 M GNRs and 5.72 ng/mL AFP. (D) Time dependence of the response for AFP with 0.80 mg/mL Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles, 1.95×10-10 M GNRs and 5.72 ng/mL AFP. All experiments were performed in PBS buffer (10.00 mM, pH 7.4).


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Interference by Coexisting Foreign Substances To determine the selectivity of the experiment for AFP under the optimum conditions, we examined the influences of various interfering substances such as metal ions, amino acids and proteins, and so on. None of the tested substances caused obvious luminescence recovery except for AFP (Figure 7). The results clearly demonstrated that the proposed method had a good selectivity for AFP detection.

Figure 7. The interference of different coexisting substances. The concentration of each substance was as follows: K+: 5.85×10-5 g/mL; Na+: 3.45×10-4 g/mL; Zn2+: 2.44×10-7 g/mL; Mg2+： 9.00×10-6 g/mL；Ca2+： 1.50×10-5 g/mL； citric acid: 7.20×10-4 g/mL; ascorbic acid ： 6.60×10-3 g/mL； glucose: 6.75×10-3 g/mL; urea： 2.25×10-5 g/mL； dopamine: 1.27×


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10-5 g/mL; immunoglobulin G (IgG): 3.75×10-4 g/mL; human serum albumin (HSA): 4.31×10-7 g/mL; arginine(Arg)： 6.53×10-5 g/mL；lysine (Lys)： 5.48×10-5 g/mL；glutathione (GSH): 2.60×10-4 g/mL ；cysteine (Cys): 4.54×10-5 g/mL; prostate specific antigen (PSA): 2.25×10-8 g/mL; ferritin in serum(SF): 2.25×10-8 g/mL; AFP: 7.62×10-9 g/mL.

Calibration Curves and Limit of Detection Figure 8 illustrates that we achieved a good linear relationship between the intensity of the recovered luminescence of the upconversion nanoparticles and the concentration of added AFP in PBS buffer solution. The linear range for AFP detection was from 0.18 ng/mL to 11.44 ng/mL. The linear regression equation: I-I0=360.94 + 154.76C, with a correlation coefficient of 0.99. The detection limit (3S/K) was defined as 0.16 ng/mL ( with S meaning standard deviation of the blank sample for ten observation and K corresponding to the slope of the linear regression equation in Figure 8).


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Figure 8. Luminescence emission spectra of the sensing system in PBS buffer solution after addition of various concentrations of AFP (A), a→g: 0 ng/mL; 0.18 ng/mL; 1.43 ng/mL; 2.86 ng/mL; 5.72 ng/mL; 7.62 ng/mL; 11.44 ng/mL. And the calibration curve for testing AFP (B) To further test the LET system in complex matrixes, the detection parameters of AFP in serum were carried out by spiking appropriate amounts of AFP in AFP depleted serum (Figure 9). The serum was diluted 100-fold by PBS buffer solution as the experimental medium. The experimental process was the same as AFP determination in PBS buffer solution. The linear range was from 0.18 ng/mL to 11.44 ng/mL and the linear regression equation: I-I0=209.32 + 124.75C, with a correlation coefficient of 0.99 and the detection limit was 0.17 ng/mL. Real serum contains many complex substances, so this system could be used for detecting AFP in serum.

Figure 9. Luminescence emission spectra for the sensing system in AFP depleted serum after addition of various concentrations of AFP (A), a→g: 0 ng/mL; 0.18 ng/mL; 1.43 ng/mL; 2.86 ng/mL; 5.72 ng/mL; 7.62 ng/mL; 11.44 ng/mL. And the calibration curve for testing AFP (B).


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Measurement of AFP in Human Serum Samples This LET method was applied for the determination of the AFP level in serum samples (provided by Yijishan Hospital of Wannan Medical College, Wuhu) after proper dilution with buffer. The experimental results are listed Table 1. The AFP levels measured in human serum samples with this LET system were in good agreement with the clinical data. The clinical data were obtained by protein chip method using a luminex-200 system.

Table1. Assay results of AFP in serum samples.

Clinical data

Proposed method R.S.D.(n=3)％

Blood samples (ng/mL)

(ng/mL)

1

5.44

5.52

1.1

2

6.37

6.52

2.0

3

146.64

145.80

1.2

4

209.61

207.81

1.6

5

274.50

280.80

2.0


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CONCLUSION In this work, we have successfully developed a new model of a luminescence energy transfer system for detecting AFP based on the stronger affinity between AFP and Ab1 than the electrostatic interaction between Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles and GNRs. NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles with a NIR emission at 804 nm were utilized as the detection signal in this study, which take advantage of the merits in NIR region.47, 48 The efficiency for LET is related to the overlap between the emission spectrum of the donor and the absorption spectrum of the acceptor. To improve the efficiency of the LET system, the GNRs with the longitudinal SPR band around 790 nm were employed as the acceptor. The results showed a highly sensitive way to detect cancer markers in clinical analysis. Therefore, this methodology may introduce a new application in the red/NIR region for other biological and clinical research.

AUTHOR INFORMATION Corresponding Author E-mail: [email protected] (H. C.); [email protected](L.W.).

ACKNOWLEDGMENT This work was financially supported by Natural Science Foundation of China (21275008, 21301008), Anhui Provincial Natural Science Foundation (1408085QB40), Special and Excellent Research Fund of Anhui Normal University.


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REFERENCES (1. Wang, L.; Yan, R.; Huo, Z.; Wang, L.; Zeng, J.; Bao, J.; Wang, X.; Peng, Q.; Li, Y. Fluorescence Resonant Energy Transfer Biosensor Based on Upconversion ‐ Luminescent Nanoparticles. Angew. Chem. Int. Ed. 2005, 44 (37), 6054-6057. 2. Haase, M.; Schäfer, H. Upconverting nanoparticles. Angew. Chem. Int. Ed. 2011, 50 (26), 5808-5829. 3. Zhou, J.; Liu, Z.; Li, F. Upconversion nanophosphors for small-animal imaging. Chem. Soc. Rev. 2012, 41 (3), 1323-1349. 4. Wang, Y.; Shen, P.; Li, C.; Wang, Y.; Liu, Z. Upconversion fluorescence resonance energy transfer based biosensor for ultrasensitive detection of matrix metalloproteinase-2 in blood. Anal. Chem. 2012, 84 (3), 1466-1473. 5. van de Rijke, F.; Zijlmans, H.; Li, S.; Vail, T.; Raap, A. K.; Niedbala, R. S.; Tanke, H. J. Up-converting phosphor reporters for nucleic acid microarrays. Nat. Biotech 2001, 19 (3), 273276. 6. Yang, J.; Shen, D.; Li, X.; Li, W.; Fang, Y.; Wei, Y.; Yao, C.; Tu, B.; Zhang, F.; Zhao, D. One-Step Hydrothermal Synthesis of Carboxyl-Functionalized Upconversion Phosphors for Bioapplications. Chem. – Eur. J. 2012, 18 (43), 13642-13650. 7. Ding, Y.; Zhu, H.; Zhang, X.; Zhu, J.-J.; Burda, C. Rhodamine B derivativefunctionalized upconversion nanoparticles for FRET-based Fe 3+-sensing. Chem. Commun 2013, 49 (71), 7797-7799. 8. Li, H.; Wang, L. NaYF4: Yb3+/Er3+ nanoparticle-based upconversion luminescence resonance energy transfer sensor for mercury (II) quantification. Analyst 2013, 138 (5), 15891595. 9. Wang, F.; Liu, X. Recent advances in the chemistry of lanthanide-doped upconversion nanocrystals. Chem. Soc. Rev. 2009, 38 (4), 976-989. 10. Wang, L.; Li, Y. Green upconversion nanocrystals for DNA detection. Chem. Commun 2006, 24, 2557-2559. 11. An, M.; Cui, J.; He, Q.; Wang, L. Down-up-conversion luminescence nanocomposites for dual-modal cell imaging. J. Mater. Chem. B. 2013, 1 (9), 1333-1339. 12. Heer, S.; Kömpe, K.; Güdel, H. U.; Haase, M. Highly Efficient Multicolour Upconversion Emission in Transparent Colloids of Lanthanide-Doped NaYF4 Nanocrystals. Adv. Mater. 2004, 16 (23-24), 2102-2105. 13. Ma, Y.; Wang, L. Upconversion luminescence nanosensor for TNT selective and labelfree quantification in the mixture of nitroaromatic explosives. Talanta 2014, 120, 100-105. 14. Wang, F.; Banerjee, D.; Liu, Y.; Chen, X.; Liu, X. Upconversion nanoparticles in biological labeling, imaging, and therapy. Analyst 2010, 135 (8), 1839-1854. 15. Chen, Z.; Chen, H.; Hu, H.; Yu, M.; Li, F.; Zhang, Q.; Zhou, Z.; Yi, T.; Huang, C. Versatile Synthesis Strategy for Carboxylic Acid−functionalized Upconverting Nanophosphors as Biological Labels. J. Am. Chem. Soc. 2008, 130 (10), 3023-3029. 16. Wang, M.; Hou, W.; Mi, C.-C.; Wang, W.-X.; Xu, Z.-R.; Teng, H.-H.; Mao, C.-B.; Xu, S.-K. Immunoassay of Goat Antihuman Immunoglobulin G Antibody Based on Luminescence Resonance Energy Transfer between Near-Infrared Responsive NaYF4:Yb, Er Upconversion Fluorescent Nanoparticles and Gold Nanoparticles. Anal. Chem. 2009, 81 (21), 8783-8789.


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Page 22 of 33

17. Ntziachristos, V.; Bremer, C.; Weissleder, R. Fluorescence imaging with near-infrared light: new technological advances that enable in vivo molecular imaging. Eur. Radio. 2003, 13 (1), 195-208. 18. Li, H.; Wang, L. Controllable Multicolor Upconversion Luminescence by Tuning the NaF Dosage. Chem. – Asian J. 2014, 9 (1), 153-157. 19. Xia, Y.; Song, L.; Zhu, C. Turn-On and Near-Infrared Fluorescent Sensing for 2,4,6Trinitrotoluene Based on Hybrid (Gold Nanorod)−(Quantum Dots) Assembly. Anal. Chem. 2011, 83 (4), 1401-1407. 20. Wang, Y.; Bao, L.; Liu, Z.; Pang, D.-W. Aptamer biosensor based on fluorescence resonance energy transfer from upconverting phosphors to carbon nanoparticles for thrombin detection in human plasma. Anal. Chem. 2011, 83 (21), 8130-8137. 21. Tu, N.; Wang, L. Surface plasmon resonance enhanced upconversion luminescence in aqueous media for TNT selective detection. Chem. Commun. 2013, 49 (56), 6319-6321. 22. Nyk, M.; Kumar, R.; Ohulchanskyy, T. Y.; Bergey, E. J.; Prasad, P. N. High contrast in vitro and in vivo photoluminescence bioimaging using near infrared to near infrared upconversion in Tm3+ and Yb3+ doped fluoride nanophosphors. Nano Lett. 2008, 8 (11), 38343838. 23. Wang, X.; Chen, J.-T.; Zhu, H.; Chen, X.; Yan, X.-P. One-Step Solvothermal Synthesis of Targetable Optomagnetic Upconversion Nanoparticles for in Vivo Bimodal Imaging. Anal. Chem. 2013, 85 (21), 10225-10231. 24. Huang, S.; Bai, M.; Wang, L. General and Facile Surface Functionalization of Hydrophobic Nanocrystals with Poly (amino acid) for Cell Luminescence Imaging. Sci. Rep. 2013, 3, 2023 - 2028. 25. Li, Z.; Zhang, Y.; Jiang, S. Multicolor core/shell‐structured upconversion fluorescent nanoparticles. Adv. Mater. 2008, 20 (24), 4765-4769. 26. Kim, J.; Piao, Y.; Hyeon, T. Multifunctional nanostructured materials for multimodal imaging, and simultaneous imaging and therapy. Chem. Soc. Rev. 2009, 38 (2), 372-390. 27. Park, Y. I.; Nam, S. H.; Kim, J. H.; Bae, Y. M.; Yoo, B.; Kim, H. M.; Jeon, K.-S.; Park, H. S.; Choi, J. S.; Lee, K. T. Comparative Study of Upconverting Nanoparticles with Various Crystal Structures, Core/Shell Structures, and Surface Characteristics. J. Phys. Chem. C 2013, 117 (5), 2239-2244. 28. Zhou, J.; Sun, Y.; Du, X.; Xiong, L.; Hu, H.; Li, F. Dual-modality in vivo imaging using rare-earth nanocrystals with near-infrared to near-infrared (NIR-to-NIR) upconversion luminescence and magnetic resonance properties. Biomaterials 2010, 31 (12), 3287-3295. 29. Chen, H.-Q.; Yuan, F.; Wang, S.-Z.; Xu, J.; Zhang, Y.-Y.; Wang, L. Near-infrared to near-infrared upconverting NaYF4:Yb3+,Tm3+ nanoparticles-aptamer-Au nanorods light resonance energy transfer system for the detection of mercuric(ii) ions in solution. Analyst 2013, 138 (8), 2392-2397. 30. Yuan, F.; Chen, H.; Xu, J.; Zhang, Y.; Wu, Y.; Wang, L. Aptamer‐Based Luminescence Energy Transfer from Near‐Infrared‐to‐Near‐Infrared Upconverting Nanoparticles to Gold Nanorods and Its Application for the Detection of Thrombin. Chem. - Eur. J. 2014. 20 (10), 2888-2894. 31. Taketa, K. α‐fetoprotein: Reevaluation in hepatology. Hepatology 1990, 12 (6), 14201432. 32. Yoshima, H.; Mizuochi, T.; Ishii, M.; Kobata, A. Structure of the asparagine-linked sugar chains of α-fetoprotein purified from human ascites fluid. Cancer Res. 1980, 40 (11), 4276-4281.


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Page 23 of 33

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33. Tang, J.; Tang, D.; Su, B.; Li, Q.; Qiu, B.; Chen, G. Specific immunoreaction-induced controlled release strategy for sensitive impedance immunoassay of a cancer marker. Analyst 2011, 136 (19), 3869-3871. 34. Chen, J.; Chen, H.; Zhou, C.; Xu, J.; Yuan, F.; Wang, L. An efficient upconversion luminescence energy transfer system for determination of trace amounts of nitrite based on NaYF4:Yb3+, Er3+ as donor. Anal. Chim. Acta. 2012, 713, 111-114. 35. Liu, C.; Wang, Z.; Wang, X.; Li, Z. Surface modification of hydrophobic NaYF4:Yb,Er upconversion nanophosphors and their applications for immunoassay. Sci. China. Chem. 2011, 54 (8), 1292-1297. 36. Dong, C.; Korinek, A.; Blasiak, B.; Tomanek, B.; van Veggel, F. C. J. M. Cation Exchange: A Facile Method To Make NaYF4:Yb,Tm-NaGdF4 Core–Shell Nanoparticles with a Thin, Tunable, and Uniform Shell. Chem. Mater. 2012, 24 (7), 1297-1305. 37. Lim, S. H.; Buchy, P.; Mardy, S.; Kang, M. S.; Yu, A. D. C. Specific Nucleic Acid Detection Using Photophysical Properties of Quantum Dot Probes. Anal. Chem. 2009, 82 (3), 886-891. 38. East, D. A.; Mulvihill, D. P.; Todd, M.; Bruce, I. J. QD-Antibody Conjugates via Carbodiimide-Mediated Coupling: A Detailed Study of the Variables Involved and a Possible New Mechanism for the Coupling Reaction under Basic Aqueous Conditions. Langmuir 2011, 27 (22), 13888-13896. 39. Huang, X.; Ren, J. Gold nanoparticles based chemiluminescent resonance energy transfer for immunoassay of alpha fetoprotein cancer marker. Anal Chim Acta 2011, 686 (1–2), 115-120. 40. Bi, S.; Zhou, H.; Zhang, S. Multilayers enzyme-coated carbon nanotubes as biolabel for ultrasensitive chemiluminescence immunoassay of cancer biomarker. Biosens. Bioelectron 2009, 24 (10), 2961-2966. 41. Wang, C.; Chen, Y.; Wang, T.; Ma, Z.; Su, Z. Biorecognition-driven self-assembly of gold nanorods: a rapid and sensitive approach toward antibody sensing. Chem. Mater. 2007, 19 (24), 5809-5811. 42. Nikoobakht, B.; El-Sayed, M. A. Preparation and Growth Mechanism of Gold Nanorods (NRs) Using Seed-Mediated Growth Method. Chem. Mater. 2003, 15 (10), 1957-1962. 43. Vetrone, F.; Boyer, J.-C.; Capobianco, J. A. Yttrium oxide nanocrystals: luminescent properties and applications. Ency. nanosci. nanotech. 2004, 10 (1), 725-765. 44. Takahashi, H.; Niidome, Y.; Niidome, T.; Kaneko, K.; Kawasaki, H.; Yamada, S. Modification of Gold Nanorods Using Phosphatidylcholine to Reduce Cytotoxicity. Langmuir 2005, 22 (1), 2-5. 45. Wang, L.; Zhang, Y.; Zhu, Y. One-pot synthesis and strong near-infrared upconversion luminescence of poly(acrylic acid)-functionalized YF3:Yb3+/Er3+ nanocrystals. Nano. Res. 2010, 3 (5), 317-325. 46. Wu, B.-Y.; Wang, H.-F.; Chen, J.-T.; Yan, X.-P. Fluorescence resonance energy transfer inhibition assay for α-fetoprotein excreted during cancer cell growth using functionalized persistent luminescence nanoparticles. J. Am. Chem. Soc. 2010, 133 (4), 686-688. 47. Kuningas, K.; Rantanen, T.; Karhunen, U.; Lövgren, T.; Soukka, T. Simultaneous Use of Time-Resolved Fluorescence and Anti-Stokes Photoluminescence in a Bioaffinity Assay. Anal. Chem. 2005, 77 (9), 2826-2834. 48. Frangioni, J. V. In vivo near-infrared fluorescence imaging. Curr. Opin. Chem. Biol. 2003, 7 (5), 626-634.


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SYNOPSIS TOC

Turn-On Detection of a Cancer Marker Based on Near-Infrared Luminescence Energy Transfer from NaYF4:Yb,Tm/ NaGdF4 Core−Shell Upconverting Nanoparticles to Gold Nanorods Hongqi Chen*, Yingying Guan, Shaozhen Wang, Yuan Ji, Mengqi Gong, Lun Wang*


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Figure 1. The TEM image (A) and EDS (B) of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles. 198x76mm (150 x 150 DPI)


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Figure 2. The TEM image of GNRs 108x112mm (150 x 150 DPI)


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Figure 3. The emission spectrum of NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (a) and the absorbance spectrum of GNRs (b). 279x215mm (150 x 150 DPI)


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Figure 4. The schematic depiction for determining AFP. 242x143mm (150 x 150 DPI)


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Figure 5. Luminescence emission spectra of (a) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles; (b) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + 7.62 ng/mL AFP; (c) Ab1modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs; (d) Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs + 2.86 ng/mL AFP; (e). Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles + GNRs + 7.62 ng/mL AFP. Conditions: Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles 0.80 mg/mL; GNRs: 1.95×10-10 M; experiments were performed in PBS buffer (10.00 mM, pH 7.4). 279x215mm (150 x 150 DPI)


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Figure 6. (A) Luminescence intensity: NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (0.80 mg/mL) + various concentrations of Ab1. (B) Luminescence quenching efficiency: Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles (0.80 mg/mL) + various concentration of GNRs. (C) The influence of different concentration of Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles on the response of AFP with 1.95×10-10 M GNRs and 5.72 ng/mL AFP. (D) Time dependence of the response for AFP with 0.80 mg/mL Ab1-modified NaYF4:Yb,Tm/NaGdF4 core−shell nanoparticles, 1.95×10-10 M GNRs and 5.72 ng/mL AFP. All experiments were performed in PBS buffer (10.00 mM, pH 7.4). 288x226mm (150 x 150 DPI)


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Figure 7. The interference of different coexisting substances. 279x215mm (150 x 150 DPI)


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Figure 8. Luminescence emission spectra of the sensing system in PBS buffer solution after addition of various concentrations of AFP 316x128mm (150 x 150 DPI)


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Figure 9. Luminescence emission spectra for the sensing system in AFP depleted serum after addition of various concentrations of AFP 308x128mm (150 x 150 DPI)


Yb,Tm - American Chemical Society

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