Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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contamination of the substances of interest (e.g., trace metals). When this strategy is used, the time needed to complete γ spectrometry before further analyses can be performed also needs to be taken into consideration. In many laboratories, the total 210 Pb concentration is determined by R spectrometry, through its daughter radionuclide 210 Po in equilibrium. The technique is simple, rapid, reliable (recoveries >95%), and shows good precision. 10,11 Furthermore, although the technique is destructive, only ca. 250 mg of dry sediment is needed, and because laboratories usually have several R detectors (they are much cheaper than Ge detectors), sample throughput is much larger than with γ spectrometry. Typical counting times range from 1 to 7 days, depending on sample activity and desired uncertainty. An experimented analyst might produce a full total 210 Pb profile within a month after sample receipt (ca. 40 samples). When R spectrometry is used, the base 210 Pb can only be indirectly determined by assuming that the 226 Ra concentration is constant along the profile and therefore estimated as the average of concentrations in the profile bottom sections, as equilibrium should have been reached. 12 However, as 226 Ra may vary along the profile, this may lead to inaccurate 210 Pbex values. It is not uncommon that, once the total 210 Pb profile is known, γ spectrometry is carried out in selected sections, thus optimizing the use of this limiting resource, although because of sample size requirements this might be impracticable for many laboratories. As γ spectrometry is carried out on the untreated sample and 210 Po on a digestate, these techniques sometimes yield results which are not fully consistent. Liquid scintillation counting (LSC), mostly used for the determination of � emitters, 13,14 has also been extensively used to quantify R emitters, such as 226 Ra, in environmental samples (mainly waters). 15,16 Some of the reported methods include direct R LSC after water sample concentration, 17 226 Ra extraction from water samples with specific scintillation cocktails such as Radaex, 18 or generation of a radium-barium sulfate coprecipitate that is transformed into a soluble chloride or nitrate. 19,20 Villa et al. 21 opted for this approach for sediment: the sediment is digested, and after the elimination of actinides as hydroxides, radium is recovered as Ra-Ba-SO4, dissolved in EDTA 0.2 M ammonia solution, and counted. However, most of these methods cannot be directly used with sediment digestates and/ or are excessively resource-consuming. (10) Sanchez-Cabeza, J. A.; Masqué, P.; Ani-Ragolta, I. J. Radioanal. Nucl. Chem. 1998, 227, 19–22. (11) Vesterbacka, P.; Ikaheimonen, T. K. Anal. Chim. Acta 2005, 545, 252– 261. (12) Binford, M. W. J. Paleolimnol. 1990, 3, 253–268. (13) Pujol, L.; Sanchez-Cabeza, J. A. J. Radioanal. Nucl. Chem. 1999, 2, 391– 398. (14) Liong Wee Kwong, L.; LaRosa, J. J.; Lee, S. H.; Povinec, P. P. J. Radioanal. Nucl. Chem. 2000, 248, 751–755. (15) Salonen, L. Sci. Total Environ. 1993, 130-131, 23–35. (16) Salonen, L.; Hukkanen, H. J. Radioanal. Nucl. Chem. 1997, 226, 67–74. (17) Sanchez-Cabeza, J. A.; Pujol, L. Analyst 1998, 123, 399–403. (18) Aupiais, J. Anal. Chim. Acta 2005, 532, 199–207. (19) Repinc, U.; Benedik, L. J. Radioanal. Nucl. Chem. 2002, 254, 181–185. (20) Galan-Lopez, M.; Martin-Sanchez, A.; Tosheva, Z.; Kies, A. In LSC 2005 Advances in Liquid Scintillation Spectrometry; Chalupnik, S., Schoenhofer, F., Noakes, J., Eds.; Radiocarbon: Tucson, AZ, 2006; pp 165-170. (21) Villa, M.; Moreno, H. P.; Manjón, G. Radiat. Meas. 2005, 39, 543–550. 6848 Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 After 210 Po analysis for R spectrometry, 226 Ra remains in solution. In this work, we propose, develop, and validate a new method to determine 226 Ra by 222 Rn emanation to a scintillation cocktail, which eliminates the need to perform any further purification, and counting with an ultralow background liquid scintillation system. EXPERIMENTAL SECTION Equipment. An ultralow background liquid scintillation system, Quantulus 1220 TM (Wallac, Turku, Finland), was used to carry out this work. In this system, background is reduced by an optimized combination of active and passive shields. A pulseshape analysis (PSA) circuit permits the discrimination of pulses produced by R and � radiation by comparing the area of the pulse tail after 50 ns from the start with its total area. Pulse-shape discrimination is accomplished using a software adjustable parameter (PSA parameter) which can vary between 1 and 256. 16,22 Quenching (sample extinction) was quantified with the standard quenching parameter (SQP(E)) which is used to determine the counting efficiency for each sample through calibration curves. 23,24 Counting was performed with Wallac OptiScint HiSafe III, a diisopropyl naphthalene based aqueous immiscible cocktail, and low-diffusion PE counting vials (Packard BioScience). Counting Solutions. The tracer solutions were prepared by gravimetrically spiking 226 Ra/2 M HNO3 (NIST, SRM4967, U.S.A.) into known amounts of deionized water contained in 20 mL low-diffusion PE counting vials. OptiScint HiSafe was then added to reach a total admixture volume of 20 mL. These were stored for 3 weeks in a dark temperature-controlled area to allow in-growth and equilibrium of the radioactive progenies. The background solutions, used for calibration purposes, were prepared with 10 mL of deionized water, acidified to match the standard solutions, to which 10 mL of the scintillation cocktail was added. In all cases, quenching was changed by adding different amounts of CCl 4, ranging from 0 to 200 µL. All reagents used in the experiments were of analytical grade (Fisher Scientific). RESULTS When counting a 226 Ra aqueous solution with an immiscible scintillant (such as OptiScint Hisafe), the R emitter 226 Ra decays to the R emitter 222 Rn (T1/2 ) 3.8332 ± 0.0008 days). Radon is highly soluble in oil-based scintillators and is selectively extracted in the cocktail, suffering some decay while this process takes place. Once solubilized in the organic phase, 222 Rn decays to the R emitter 218 Po (T1/2 ) 3.094 ± 0.006 min), this mainly decays to the � emitter 214 Pb (T1/2 ) 26.8 ± 9 min), which decays to the � emitter 214 Bi (T1/2 ) 19.9 ± 0.4 min), which mainly decays to the R emitter 214 Po (T1/2 ) 162.3 ± 1.2 µs), and this one to 210 Pb (T1/2 ) 22.23 ± 0.12 years). Therefore, in the scintillant, and after an appropriate equilibration time (usually set to about 3 weeks), the R emitters 222 Rn, 218 Pb, and 214 Pb are in secular equilibrium (Figure 1). As the probability of these R decays is in all cases close to one, the maximum (22) Kaihola, L. J. Radioanal. Nucl. Chem. 2000, 243, 313–317. (23) Villa, M.; Manjon, G.; Garcia-Leon, M. Nucl. Instrum. Methods Phys. Res., Sect. A 2003, 496 (2-3), 413–424. (24) Sanchez-Cabeza, J. A.; Pujol, L. Health Phys. 1995, 68 (5), 674–82.
Figure 1. 226 Ra spectrum by liquid scintillation (only 6 × 10 -4 Bq). Vertical dashed lines indicate the approximate counting windows. efficiency reached when counting all R events should be 300%. This is another advantage of R versus γ spectrometry, where final counting efficiencies for 226 Ra, depending on the counting configuration, normally do not usually exceed a few percent. The magnitudes used for optimization were those related to counting precision. • Counting efficiency E: this is the ratio between the observed number of counts and the 226 Ra decays. The maximum observed efficiency is close to 300% (Figure 1) because with this method we simultaneously count R particles from 222 Rn, 218 Pb, and 214 Pb in equilibrium. • Minimum detectable activity (MDA): 25 we used the following expression 2.71 + 4.65√Bt MDA) tVE where B is the background count rate, t is the counting time, and V is the volume of solution used (in the case of sediment analysis, V was substituted by m, the mass of the aliquot analyzed). • Figure of merit: this is a magnitude commonly used to compare methods, which emphasizes counting efficiency (and therefore sample throughput). FM was calculated as FM ) E 2 /B. In this section, we describe the results of the optimization process for the relevant counting parameters. Optimal Admixture Composition. Once the scintillation vial and total volume were fixed, the optimal “scintillator-to-water” ratio was optimized. We prepared and counted several composition mixtures with tracer solutions and background solutions, by using 1, 5, 10, 15, and 19 mL of scintillant (Figure 2). Not unexpectedly, the highest efficiency was observed for the maximum volume of scintillant, as this maximizes radiation interaction with the detector (the scintillant). However, as the scintillant itself is an important source of background, this also increases the MDA value. This behavior is well-captured with the FM, which shows a maximum value for a 10:10 mL admixture. Therefore, we used this proportion in all experiments. Optimal PSA Parameter. The PSA circuit of Quantulus 1220 sends the signals to one of the two multichannel analyzers Figure 2. Determination of the optimal admixture composition. Figure 3. Change of background and 226 Ra counting efficiency with PSA. depending on the result of the PSA analysis. However, this method is not error-free and shows interference as (i) R and � events can be wrongly assigned and (ii) the efficiency of this method depends on the energy of the incident radiation particles. 26 In order to minimize the interference, tracer solutions of 226 Ra activity 5.34 Bq and background solutions, with a 10:10 mL admixture composition, were counted by LSC during 12 h and with the PSA parameter ranging from PSA ) 0 (no R-� discrimination) to PSA ) 256 (maximum R-� discrimination) with a step increment of 5 (Figure 3). The background signals assigned to the R spectrum have a varied nature and may include γ interactions with the scintillant, � events wrongly assigned to the R spectrum, R emission from impurities in the vial, and a variety of cosmic-ray originated signals. Therefore, the background spectrum does not show prominent peaks. The background count rate versus PSA plot (Figure 3) shows an almost constant value until PSA ∼ 90, as all events are recorded in the R spectrum, and then its value decays smoothly to a value close to 0 cpm, when all background events are recorded in the � spectrum. On the other hand, the 226 Ra spectrum shows well-resolved peaks within a quite narrow energy window (Figure 1), and the effect of the R particle energy on the interference is clearly shown in the efficiency versus PSA curve (Figure 3). Until PSA ∼ 110, the total efficiency exceeds the maximum value of 300% because of the misclassification of background and � events in the R spectrum. In the PSA range of 120-160, most background (25) Currie, L. A. Anal. Chem. 1968, 40, 586–593. (26) Pujol, L.; Sanchez-Cabeza, J. A. Analyst 1997, 122, 383–385. Analytical Chemistry, Vol. 82, No. 16, August 15, 2010 6849
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Anal. Chem. 2010, 82, 6745-6750 Let
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purchased from Invitrogen-Molecular
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Figure 3. Electropherograms of TPP-
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Anal. Chem. 2010, 82, 6751-6755 Res
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(pH 8.0) with cysteine and cystamin
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Figure 4. Ion mobility mass spectra
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GOx glucose + O298 gluconic acid +
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coater at 2000 rpm for 20 s, and th
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Figure 5. Comparison of cyclic volt
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in channels with either no grooves
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indicators of atmospheric processin
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Figure 1. GC/MS total ion chromatog
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Table 2. Concentrations and Stable
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on a substrate are preferred. 20-24
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Figure 4. SERS analysis of NAADP co
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Anal. Chem. 2010, 82, 6775-6781 Hig
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tion, 2 µL of proprionaldehyde wer
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Figure 4. Analysis of 2a by HPLC-MS
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eaction of 1a with PBH can be condu
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Numerous references had demonstrate
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Figure 1. TEM images of the prepare
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Figure 4. Schematic representation
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esult in a big SPR signal change wi
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also reduces chemical noise, which
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Table 1. Extraction Yields, Liquid
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scanning of AMPP amides of the anal
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Page 101 and 102: Figure 5. Quantitative calibration
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Page 125 and 126: ment. Therefore, the long-time drea
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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),
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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
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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
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Anal. Chem. 2010, 82, 7049-7052 Dev
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Figure 2. Comparative analysis of d
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