Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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Figure 3. AFM images of 14.51 nm Fe3O4 MNP (A) and 32.82 nm Fe3O4 MNP (B) modified SPR gold substrate. The concentration of Fe3O4 MNPs is 1.6 nM, and the assembly time is 10 min. coverage of Fe3O4 MNPs on SPR gold film could be reached at a higher concentration due to the rapid diffusion adsorption. It should be pointed out that the SPR angle shift resulting from the adsorption of Fe3O4 MNPs is much higher than that of the value resulting from the adsorption of most biomolecules under the same concentration. 40,41 It means Fe3O4 MNPs greatly enhance the signal of SPR spectroscopy. Importantly, the SPR angle shift could be further increased when 32.82 nm Fe3O4 MNPs are used. Figure 2B shows the SPR angle shift curves resulting from the adsorption of 32.82 nm Fe3O4 MNPs with the same concentration sequence as 14.51 nm Fe3O4 MNPs. With the increase of Fe3O4 MNP concentrations from 0.016 to 1.6 nM, the SPR angle shifts increase from 79.22 to 2479.79 m°. To further understand the size effect of Fe3O4 MNPs on the SPR response, the relation between SPR angle shifts and the concentrations of Fe3O4 MNPs with the two sizes are compared in Figure 2C. Obviously, much larger angle shifts are observed for 32.82 nm Fe3O4 MNPs because of its larger moleculer weight. Besides that, the affinity constants and surface coverage for both kinds of Fe3O4 MNPs could also be obtained from Figure 2C. From the data of Figure 2C, we can see that the plasmon resonance shifts with the increasing Fe3O4 MNP concentration and a simple Langmuir isotherm 42 could be used to fit the data. The Langmuir equation used to fit the data is given by KA [C] Fe3O4 ∆λ ) ∆λmax1 + KA [C] Fe3O4 Where ∆λ is the angle shift caused by the adsorption of Fe3O4 MNPs, ∆λmax is the angle shift which will be observed at saturation, KA is the apparent equilibrium affinity constant, and [C]Fe3O4 is the concentration of Fe3O4 MNPs. Using the above equation, KA values for 14.51 and 32.82 nm Fe3O4 MNPs are (40) Mullett, W. M.; Lai, E. P. C.; Yeung, J. M. Methods 2000, 22, 77–91. (41) Hoa, X. D.; Kirk, A. G.; Tabrizian, M. Biosens. Bioelectron. 2007, 23, 151– 160. (42) Lee, H. J.; Wark, A. W.; Corn, R. M. Langmuir 2006, 22, 5241–5250. 6786 Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 (1) calculated to be around 7.14 × 10 8 and 2.63 × 10 9 M -1 , respectively, which are similar to the value of NPs as found in literature. 43 The saturation responses ∆λmax of 14.51 and 32.82 nm are calculated to be 2165 and 3134 0 m, respectively. It can be seen that 32.82 nm Fe3O4 MNPs have slightly higher affinity for adsorption on the sensing surface due to their larger molecular weight; therefore, they show larger angle shifts as well as higher saturation response (∆λmax) compared to 14.51 nm Fe3O4 MNPs. In our following experiments, we, therefore, use 32.82 nm Fe3O4 MNPs. The ratio (∆λ)/(∆λmax) gives the fraction of surface coverage, and if the bulk concentration of [C]Fe3O4 is equal to (1)/(KA), half of the surface sites will be occupied ((∆λ)/(∆λmax) ) 0.5). On the basis of the experimental results and the fitting parameters, we calculate that 80% of the coverage ((∆λ)/(∆λmax) ) 0.8) will be obtained when 32.82 nm Fe3O4 MNPs with a concentration of 1.6 nM are used. This concentration will be used in our following experiments because the changes in SPR angle shift (∆λ) will be very small even if we further increase the concentration of Fe3O4 MNPs. To demonstrate that a dense monolayer of MNPs could be formed on a SPR substrate within 10 min using a 1.6 nM Fe3O4 MNP solution as assembly solution, AFM is used to evaluate the surface morphology of MNP modified SPR gold substrate. Figure 3A,B shows the AFM images of 14.51 nm Fe3O4 MNPs and 32.82 nm Fe3O4 MNPs on SPR gold substrate, respectively. As seen from the AFM figures, the nanoislands are formed after MNPs are assembled. The diameter of nanoislands in Figure 3A is smaller than that of the value in Figure 3B due to different diameter of MNPs. However, the dense layer of MNPs on SPR gold substrate has been observed from Figure 3 for both kinds of Fe3O4 MNPs at the present experimental condition, which indicates the adsorption of Fe3O4 MNPs on SPR gold substrate is fast and the present experimental condition is suitable for further experiments. (43) Liao, W. S.; Chen, X.; Yang, T. L.; Castellana, E. T.; Chen, J. X.; Cremer, P. S. Biointerphases 2009, 4, 80–85.
Figure 4. Schematic representation of the SPR biosensor for the detection of the small molecules. Fe3O4 MNP Enhanced SPR Sensing for the Detection of Small Molecules. To further demonstrate the practicability of the amplification effect of Fe3O4 MNPs in an enhancing SPRbased bioassay, a novel SPR sensor based on indirect competitive inhibition assay (ICIA) for the detection of adenosine is constructed. The principle of this SPR sensor is shown in Figure 4. The partial complementary thiolated ss-DNA of antiadenosine aptamer is first immobilized on SPR gold film as a sensing surface. When Fe3O4 MNP-antiadenosine aptamer conjugate solution is added to the SPR cell in the absence of adenosine, Fe3O4 MNP-antiadenosine aptamer conjugates will be adsorbed to the SPR sensor by the DNA hybridization reaction and result in a huge change of SPR signal due to the amplification effect of Fe3O4 MNPs. However, the change of SPR signal will decrease after Fe3O4 MNP-antiadenosine aptamer conjugates bind with adenosine. This is because adenosine reacts with antiadenosine aptamer in Fe3O4 MNPantiadenosine aptamer conjugates and changes its structure from ss-DNA to tertiary structure, which cannot hybridize with its partial complementary ss-DNA immobilized on the SPR gold surface. Thus, the change of SPR signal will decrease with the increase of the number of Fe3O4 MNP-antiadenosine aptamer conjugates possessing tertiary structure, which is proportional to the concentration of adenosine. The essential prerequisite of this detection is to prepare Fe3O4 MNP-antiadenosine aptamer conjugates. For this purpose, EDC and NHS are used as a carboxyl activating agent for the coupling of primary amines in aptamer and carboxyl groups in polymer coated MNPs. To demonstrate that MNPs have been labeled by an aptamer successfully, FT-IR spectroscopy is used to evaluate the surface modification of MNPs before and after aptamer labeling. Figure 5 shows the IR absorbance spectra of Fe3O4 MNPs before and after the aptamer label. For the polymer coated Fe3O4 MNPs, the absorbance peak near 1710 cm -1 is assigned to υ (CdO) (stretch vibration of carbonyl) and amide I, peaks at 1550 cm -1 are assigned to amide II, and the peaks at 2854 and 2925 cm -1 are assigned to υs (C-H2) and υas (C-H2) (symmetric and asymmetric stretch vibrations of C-H2), respectively. In the region of 3200-3570 Figure 5. FT-IR of polymer coated Fe3O4 MNPs before and after aptamer labeling. cm -1 , a peak due to the O-H stretch is expected. Several new peaks are observed after MNPs are labeled by the aptamer. The peaks at 1048 and 1211 cm -1 are attributed to stretching vibrations of the PO 2- in the aptamer. It should be pointed out that we are not able to confirm the presence of DNA-related peaks at 1550 cm -1 because the peak overlaps with amide II. After Fe3O4 MNP-antiadenosine aptamer conjugates are prepared successfully, the SPR detection is carried out according to the principle described in Figure 4. Figure 6A shows the SPR angle-time curves of the separation products obtained after Fe3O4 MNP-antiadenosine aptamer conjugates are reacted with different concentrations of adenosine for 30 min. In the absence of adenosine, Fe3O4MNP-antiadenosine aptamer conjugates in solution directly hybridize with ss- DNA immobilized on SPR gold film, resulting in the largest SPR angle shift (∼1082.94 m°). This angle shift is much larger than that of the angle shift resulting from the binding of ss-DNA or most of the protein. The SPR angle shift resulting from the binding of Fe3O4 MNP-antiadenosine aptamer conjugates decreases (∼820.14 m°) after Fe3O4 MNP-antiadenosine aptamer conjugates are reacted with Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 6787
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 and 40: Numerous references had demonstrate
Page 41: Figure 1. TEM images of the prepare
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
Page 91 and 92: 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
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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
Page 163 and 164:
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
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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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