Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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mL of Fe3O4 MNP solution under stirring for 0.5 h to activate the carboxyl group on the surface of Fe3O4 MNPs. After that, 100 µL of 37.7 µM amino-modified antiadenosine aptamer was added in this solution, and they were allowed to react for 2 h to immobilize aptamer on the surface of Fe3O4 MNPs. After that, 1 M ethanolamine was added for 1 h to block the unreacted carboxyl groups. Then, this solution was centrifuged at 14 000g at room temperature for 25 min twice to remove the free amino-aptamer. At last, the Fe3O4 MNPs were dispersed in 5 mL of pH 8.0 PBS buffer and stored at 4 °C. In Situ SPR Measurement. The SPR experiments were done using Eco Chemie Autolab SPR systems (Brinkmann Instruments, New York). 29,30 It works with a laser diode fixed at a wavelength of 670 nm, using a vibrating mirror to modulate the angle of incidence of the p-polarized light beam on the SPR substrate. The instrument was equipped with a cuvette. A gold sensor disk (25 mm in diameter) was mounted on the hemicylindrical lens (with index-matching oil) to form the base of the cuvette. The cuvette could contain sample with adjustable volume from 10 to 1000 µL. An O-ring (3 mm inner diameter) between the cuvette and disk prevents leakage. An autosampler (Eco Chemie) with a controllable aspirating-dispensing-mixing pipet was used to add samples into the cuvette and provide constant mixture by aspiration and dispensing during measurements. This experimental arrangement maintains a homogeneous solution and reproducible hydrodynamic conditions. The injection rate and mixing rate for all samples were 10 and 40 µL/s, respectively, with the total volume for all samples dispensed in the SPR cell equal to 40 µL. This setup allows us to measure the SPR angle shift in millidegrees (m°) as a response unit to quantify the binding amount of macromolecules to the sensor surface. Details of the experiment are as follows. The SPR gold film was initially immersed into the thiolated ss-DNA solution for 12 h in order to assemble the monolayer of ss-DNA. Then, the modified gold film was thoroughly rinsed with 50 mM Tris-HCl buffer and water to remove the weakly adsorbed ss-DNA. Then, the ss- DNA modified SPR gold film was immersed in 100 µM 6-mercaptohexanol for 1htoblock the uncovered gold surface. This gold film was used as a sensing surface to detect the amount of aptamer possessing ss-DNA structure on Fe 3O4 MNP-aptamer conjugates, which can be adjusted by the concentration of adenosine added to the Fe3O4 MNP-aptamer conjugate solution. The detection procedure is made up of two steps. First, Fe3O4 MNP-aptamer conjugate solution was mixed with different concentrations of adenosine for 30 min. After that, adenosine bound with Fe3O4 MNP-aptamer conjugates were separated and enriched by centrifuging at 14 000g for 1 h twice. The precipitation was dispersed in PBS again. The resulting solution was injected into the SPR cell, and the SPR angle-time curve was recorded. In order to reduce the disturbance of DNA denaturation that resulted from the regenerating process for the detecting results, we change a new substrate after each detection. The modification of each gold substrate is carried out under the same experimental condition. To confirm the reproducibility of the detection result, different concentrations (29) Wang, J. L.; Zhou, H. S. Anal. Chem. 2008, 80, 7174–7178. (30) Wang, J. L.; Munir, A.; Li, Z. H.; Zhou, H. S. Biosens. Bioelectron. 2009, 25, 124–129. 6784 Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 of the MNPs and adenosine are repeatedly detected for three times. RESULTS AND DISCUSSION Characterization of Fe3O4 MNPs. Although soluble Fe3O4 MNPs could be easily synthesized by coprecipitation of aqueous Fe 2+ /Fe 3+ salt solutions with the addition of a base under an inert atmosphere at room temperature or at elevated temperature, the size, shape, and composition of the MNPs very much depends on the type of salts used (e.g., chlorides, sulfates, nitrates), the Fe 2+ /Fe 3+ ratio, the reaction temperature, the pH value, and ionic strength of the media. 31-33 Furthermore, the Fe3O4 MNPs are also easy to aggregate. Inspired by the synthesis of high-quality semiconductor nanocrystals and oxides in nonaqueous media by thermal decomposition, 34-36 monodisperse Fe3O4 MNPs with controlled size has essentially been synthesized through thermal decomposition of ion compounds in high-boiling organic solvents. 37,38 Here, we synthesize Fe3O4 MNPs by the pyrolysis of iron carboxylate in an organic phase. By changing the ratio of FeO(OH) and oleic acid, two kinds of Fe3O4 MNPs are prepared. Figure 1A,B provides the TEM images of the prepared Fe3O4 MNPs and their size distribution. The average size of Fe3O4 MNPs derived from Figure 1A,B are 14.51 nm (n ) 300 particles) and 32.82 nm (n ) 138 particles), respectively. Importantly, both kinds of Fe3O4 MNP size distributions are narrow, which indicates the prepared Fe3O4 MNPs are monodisperse. After transferring Fe3O4 MNPs from organic reagent to water solution by the amphiphilic polymer, Fe3O4 MNPs show a good stability in both PBS and Tris buffer due to the large hydrodynamic size of polymer (data not shown), which are all beneficial for the acquisition of accurate and repeated SPR analytical results. SPR Response and Concentration Dependence of Fe3O4 MNPs. The basis of a particle-enhanced bioassay is that biomolecular interaction events lead to particle immobilization, i.e., more immobilized proteins yield higher particle coverage. 39 Currently, most SPR instruments are able to quantitatively detect the concentration of biomolecules through calculating the SPR angle shift enhanced by the binding of nanoparticle. Considering our synthesized Fe3O4 MNPs are protected by negative polymer, 2-mercaptoethyamine is used on SPR Au substrates for the adsorption of Fe3O4 MNPs. The thiol group binds to the Au surface, leaving the amine group free to bind with carboxyl group in soluble Fe3O4 MNPs by electrostatic interaction. (31) Lu, A. H.; Salabas, E. L.; Schuth, F. Angew. Chem., Int. Ed. 2007, 46, 1222– 1244. (32) Kang, Y. S.; Risbud, S.; Rabolt, J. F.; Stroeve, P. Chem. Mater. 1996, 8, 2209–2211. (33) Massart, R. IEEE Trans. Magn. 1981, 17, 1247–1248. (34) Murray, C. B.; Norris, D. J.; Bawendi, M. G. J. Am. Chem. Soc. 1993, 115, 8706–8715. (35) Peng, X.; Wickham, J.; Alivisatos, A. P. J. Am. Chem. Soc. 1998, 120, 5343– 5344. (36) O’Brien, S.; Brus, L.; Murray, C. B. J. Am. Chem. Soc. 2001, 123, 12085– 12086. (37) Sun, S.; Zeng, H.; Robinson, D. B.; Raoux, S.; Rice, P. M.; Wang, S. X.; Li, G. J. Am. Chem. Soc. 2004, 126, 273–279. (38) Redl, F. X.; Black, C. T.; Papaefthymiou, G. C.; Sandstrom, R. L.; Yin, M.; Zeng, H.; Murray, C. B.; O’Brien, S. P. J. Am. Chem. Soc. 2004, 126, 14583– 14599. (39) Lyon, L. A.; Pena, D. J.; Natan, M. J. J. Phys. Chem. B 1999, 103, 5826– 5831.
Figure 1. TEM images of the prepared Fe3O4 MNPs: (A) 14.51 nm and (B) 32.82 nm. Figure 2. Variation of SPR angle-time curves with the concentration of Fe3O4 MNPs: (A) 14.51 nm; (B) 32.82 nm. (C) Comparison of the variation of SPR angle shift with the concentration of Fe3O4 MNPs of 14.51 and 32.82 nm, respectively. Figure 2 illustrates the changes that occur in the SPR angle shift as a function of the concentration for 14.51 nm (Figure 2A) and 32.82 nm (Figure 2B) Fe3O4 MNPs. In the case of 14.51 nm Fe3O4 MNPs (Figure 2A), the SPR angle shifts gradually increase from 6.4 to 1111.06 m° with the increase of Fe3O4 MNP concentrations from 0.016 to 1.6 nM. It is evident that a higher Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 6785
Page 1 and 2: Anal. Chem. 2010, 82, 6745-6750 Let
Page 3 and 4: purchased from Invitrogen-Molecular
Page 5 and 6: Figure 3. Electropherograms of TPP-
Page 7 and 8: Anal. Chem. 2010, 82, 6751-6755 Res
Page 9 and 10: (pH 8.0) with cysteine and cystamin
Page 11 and 12: Figure 4. Ion mobility mass spectra
Page 13 and 14: GOx glucose + O298 gluconic acid +
Page 15 and 16: coater at 2000 rpm for 20 s, and th
Page 17 and 18: Figure 5. Comparison of cyclic volt
Page 19 and 20: in channels with either no grooves
Page 21 and 22: indicators of atmospheric processin
Page 23 and 24: Figure 1. GC/MS total ion chromatog
Page 25 and 26: Table 2. Concentrations and Stable
Page 27 and 28: on a substrate are preferred. 20-24
Page 29 and 30: Figure 4. SERS analysis of NAADP co
Page 31 and 32: Anal. Chem. 2010, 82, 6775-6781 Hig
Page 33 and 34: tion, 2 µL of proprionaldehyde wer
Page 35 and 36: Figure 4. Analysis of 2a by HPLC-MS
Page 37 and 38: eaction of 1a with PBH can be condu
Page 39: Numerous references had demonstrate
Page 43 and 44: Figure 4. Schematic representation
Page 45 and 46: esult in a big SPR signal change wi
Page 47 and 48: also reduces chemical noise, which
Page 49 and 50: Table 1. Extraction Yields, Liquid
Page 51 and 52: scanning of AMPP amides of the anal
Page 53 and 54: Anal. Chem. 2010, 82, 6797-6806 δ
Page 55 and 56: Figure 1. Schematic view of the pre
Page 57 and 58: from the specimen and enclosed in a
Page 59 and 60: Figure 4. (A) IRMS mass-44 chromato
Page 61 and 62: Table 3. Ambient Measurement Result
Page 63 and 64: Anal. Chem. 2010, 82, 6807-6813 Dir
Page 65 and 66: Polymerase (1 U per sample). Reacti
Page 67 and 68: of which was constant for all ampli
Page 69 and 70: analytically useful signals at less
Page 71 and 72: carbon black and RP-C18 for the ext
Page 73 and 74: solution in an equal volume, and 1
Page 75 and 76: Table 2. Concentrations and Ratios
Page 77 and 78: Anal. Chem. 2010, 82, 6821-6829 Mac
Page 79 and 80: mg/mL protein, followed by separati
Page 81 and 82: Figure 2. Productivity of SEQUST an
Page 83 and 84: Figure 4. High-resolution MS/MS spe
Page 85 and 86: Figure 6. Characterization of PSMs
Page 87 and 88: containing T-T mismatches. 23 Based
Page 89 and 90: Figure 1. Extinction spectra of sol
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Figure 3. (A) The value of Ex650 nm
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Figure 5. Extinction spectra and co
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wished to explore the dehydration o
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constant medium for separation; we
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Figure 2. Standards of (Pi)n, n ) 1
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Figure 5. Quantitative calibration
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Anal. Chem. 2010, 82, 6847-6853 Met
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Figure 1. 226 Ra spectrum by liquid
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Table 1. Counting Properties and De
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Table 3. Analysis of 226 Ra in Sedi
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mercial microarray scanner and fabr
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Figure 2. Optical transmission meas
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Figure 5. Volcano plots detailing t
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data as well. The 41 genes in Table
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corresponding compound if its chemi
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Figure 1. Schematic illustration of
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Table 1. Absolute Quantification Re
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ment. Therefore, the long-time drea
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Figure 1. Chemically actuated micro
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Figure 2. Influence of a surfactant
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Figure 5. Device to eject and mix s
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Anal. Chem. 2010, 82, 6877-6886 Imm
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NaCl, phosphate buffer saline (PBS)
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Figure 2. Product ion mass spectra
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-70 °C resolved the problem, givin
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Table 1. Intraday Precision and Acc
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Anal. Chem. 2010, 82, 6887-6894 Fer
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Figure 1. Infrared spectra of (A) u
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Figure 3. Cyclic voltammograms obta
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Figure 6. Calculated charge from ch
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Anal. Chem. 2010, 82, 6895-6903 Ele
Page 153 and 154:
Table 1. Chemical Structure, pKa Va
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pH with a tilted baseline (Figure 1
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Table 2. Linearity and Detection Li
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ascorbic acid (AA), uric acid (UA),
Page 161 and 162:
the multielement capabilities, the
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RESULTS AND DISCUSSION Sulfur Detec
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Table 2. Molecular Properties and C
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Anal. Chem. 2010, 82, 6911-6918 Dir
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dilution and hybridization buffer.
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solution under appropriate incubati
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Figure 4. Standardization curve for
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Anal. Chem. 2010, 82, 6919-6925 Ele
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the ×10 objective, to have a large
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Figure 2. With a suitable removal o
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the NB signal in a much better foot
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Scheme 1. Reactions of Selenium Rea
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Figure 2. (a) ESI-MS spectrum showi
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Figure 4. (a) ESI-MS spectrum showi
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Anal. Chem. 2010, 82, 6933-6939 Dif
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trode 28 by a finite element using
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Figure 4. Comparison between simula
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Figure 6. Comparison between simula
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educed in the vicinity of double bo
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Figure 2. Normalized product ion ab
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Figure 4. EID (a) and IRMPD (b) of
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Anal. Chem. 2010, 82, 6947-6957 Ide
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Figure 1. Schematic flowchart showi
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difference, ppm compound Table 1. I
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isoforms, its successful use, in th
Page 211 and 212:
Figure 5. Extracted ion current ESI
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m/z 1172.935 was observed for Ser14
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linked products via affinity tags.
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Scheme 2. Fragmentation Mechanism o
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Figure 1. (A) ESI-LTQ-CID-MS 2 prod
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Figure 3. (A) ESI-LTQ-CID-MS 2 prod
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Figure 5. (A) MALDI-TOF/TOF product
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Anal. Chem. 2010, 82, 6969-6975 Ana
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Figure 3. Equilibrium response as a
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Figure 6. The average measured resp
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for the fill time, and we find that
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nucleotide tails. 3-5 Thus, the amo
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allow the use of higher aptamer con
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quent ligation of the aptamers afte
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Anal. Chem. 2010, 82, 6983-6990 Imp
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Figure 2. Configuration editor wind
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Figure 4. Dependencies between volu
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Since the time for liquid expulsion
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Anal. Chem. 2010, 82, 6991-6999 Int
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Figure 1. Representation of the Car
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Table 2. Capillary Electrophoresis
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Figure 4. (A) Typical raw electroph
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field. DNA profiles were delivered
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Another approach to handle this lim
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(principal components, PCs) in whic
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Figure 5. Relevant score plots and
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CONCLUSIONS In this paper we have i
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electroactive-species loaded liposo
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Figure 2. Qdot-based FLFTS response
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Figure 6. Fluorescence imaging of Q
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Anal. Chem. 2010, 82, 7015-7020 Sel
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Table 1. Effect of 1 D-LC Condition
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Figure 3. Peak-production rate vers
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Anal. Chem. 2010, 82, 7021-7026 Cel
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luciferase (T7 control vector) was
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Figure 3. Inhibitory effects of luc
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Anal. Chem. 2010, 82, 7027-7034 Pat
Page 285 and 286:
Electrochemistry. Electrochemical m
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Figure 2. SEM images of Au substrat
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Table 3. Film Thickness Measurement
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Anal. Chem. 2010, 82, 7035-7043 Qua
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Figure 1. (A) FL spectra of BSPOTPE
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Figure 4. (A) Variation in the FL i
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Figure 6. (A) FL spectra of BSPOTPE
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Scheme 1. Proposed Mechanism for Fl
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one or more drawbacks including poo
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Figure 2. Free fluorophores do not
Page 305 and 306:
Anal. Chem. 2010, 82, 7049-7052 Dev
Page 307 and 308:
Figure 2. Comparative analysis of d
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Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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