Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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Figure 2. SALDI-PSD MS spectra of TMAE-VLCFAs. the Supporting Information (Figures S-2 and S-3, respectively). The SALDI-TOFMS spectra for the enrichment and SALDI analysis of TMAE-VLCFAs with various MWCNTs are depicted in Figure 1A-D and compared with the MALDI-MS with CHCA as the matrix (Figure 1E). The TMAE-VLCFAs (1 µg/mL) ions were only detected in MWCNT 636843 (Figure 1A). The TMAE- VLCFAs ions were [C20:0-TMAE] + at m/z 398.4, [C22:0-TMAE] + at m/z 426.4, [C24:0-TMAE] + at m/z 454.5, and [C26:0-TMAE] + at m/z 482.5. SALDI-MS spectra showed LODs of 0.5-1 µg/ mL for MWCNT 636843, 1-5 µg/mL for MWCNT 636495 and MWCNT 636509, and 5-10 µg/mL for MWCNT 636819 (Figure 1 and S-4 in the Supporting Information). The MALDI- MS analysis with CHCA as the matrix showed an LOD of 1-5 µg/mL. In addition, more background peaks were observed in MALDI-MS spectra compared to MWCNT-based SALDI-TOFMS spectra. There were at least two factors that contributed to the analytical sensitivity in this study: the adsorption efficiency in the enrichment procedure and the desorption/ionization efficiency in SALDI. The adsorption efficiency of the MWCNTs was analyzed by HPLC/ESI-MS. The LC/MS chromatograms of TMAE-VLCFAs are shown in Figure S-5 in the Supporting Information. The calibration and validation data of the LC/MS method are given in Figure S-6 and Table S-1 in the Supporting Information. The adsorption efficiencies of the four MWCNTs for VLCFAs were >99.8%. There were no significant differences (p > 0.05) between the four MWCNTs. We attribute the varied desorption/ionization efficiency in SALDI to the different sizes of MWCNTs. The MWCNT 636843 had a much larger outer diameter (40-70 nm), a wider wall thickness (0-65 nm), and a shorter length (0.5-2 µm) than the MWCNT 636819, which had a much smaller outer diameter (
Table 2. Concentrations and Ratios of VLCFAs in Plasma from Patients with ALD and from Normal Controls as Measured by SALDI-TOFMS Postsource Decay (PSD) of TMAE-VLCFAs. The PSD MS spectra of the derivatized VLCFAs in SALDI-TOFMS, which detected a neutral loss of trimethylamine (59 Da), were similar to those in LC-MS/MS. 30 SALDI-PSD MS spectra of TMAE- VLCFAs are recorded in Figure 2. Fragment ions were detected at m/z 339.3 for C20:0, m/z 367.4 for C22:0, m/z 395.4 for C24:0, and m/z 423.4 for C26:0. Those data demonstrate that derivatized VLCFAs not only increased the ionization efficiency in SALDI but also made the PSD easier in TOFMS. Furthermore, the neutral loss fragmentations helped confirm the presence of VLCFAs in the complex plasma matrix. Quantitative Analysis of the TMAE-VLCFAs. CNT-based SALDI has been shown to have excellent reproducibility of spectrum signals, making it an acceptable modality for quantitative analysis. 22,19,32 In addition, MALDI-MS with and without the stable isotope-labeled method has been shown to be suitable for quantifiation of small compounds in biologic samples. 38-40 In this study, we used stable isotope-labeled VLCFAs (C26:0-d4, C22:0d3, C20:0-d3) as the internal standards. The mass difference of 3-4 Da between VLCFAs and isotope-labeled VLCFAs provided sufficient m/z separation space to avoid interference with isotopologues of the ions. Figure 3 shows the SALDI-TOF mass spectra of the four derivative VLCFAs. Three of the derivative isotope-labeled VLCFAs had a molar ratio of 2:1 and a peak intensity ratio of 2:1. The isotope-labeled C22:0-d3 and C26:0-d4 were both evaluated in the quantification of C24:0 as the internal standards, with C26:0-d4 showing superior linearity (R 2 regression coefficient varied from 0.9876 to 0.9969). The accuracy of the method was measured by determining the mean concentration at various concentrations of analyte and was calculated as percentage error of theoretical versus measured concentrations. Both the intra- and interday accuracy and precision of the method were determined by triplicate analysis of standard samples containing VLCFAs at the concentrations used to construct the calibration curves. Precision was estimated as the coefficient of variance (CV) of the analyses. The interday accuracy ranged from -5.41 to 3.23%, and the intraday accuracy ranged from -3.95 to 4.27% (Table C24:0/C22:0 (ion peaks intensity ratio) C24:0/C22:0 (conc ratio) C20:0 (µg/mL) C22:0 (µg/mL) C24:0 (µg/mL) 1 16.23 15.21 patients with ALD 17.58 1.02 1.16 2 17.25 11.81 22.21 1.15 1.88 3 14.53 9.10 15.30 1.49 1.68 4 17.89 11.47 14.70 1.19 1.28 mean ± SD 16.47 ± 1.46 11.90 ± 2.5 17.45 ± 3.41 1.21 ± 0.20 1.50 ± 0.34 GC/MS median a (5-90% range) 20.8 (11.8-29.5) 13.7 (7.0-21.1) 1.50 (1.18-1.74) 1 22.52 28.32 control group 19.52 0.64 0.69 2 20.28 26.30 20.95 0.76 0.80 3 18.86 22.13 18.25 0.76 0.82 4 18.16 18.45 14.96 0.74 0.81 mean ± SD 19.95 ± 1.92 23.80 ± 4.40 18.42 ± 2.55 0.72 ± 0.06 0.78 ± 0.06 GC/MS median a (5-90% range) 25.3 (10.5-51.0) 17.4 (8.5-35.7) 0.73 (0.48-0.89) a Value reported by Schutgens. 27 S-2 in the Supporting Information). The interassay precision ranged from 1.05 to 7.52% and the intra-assay precision ranged from 0.89 to 8.85% (Table S-2 in the Supporting Information). It was found that the quantification of VLCFAs by MWCNT-based SALDI, using isotope-labeled VLCFAs as internal standards, was of acceptable accuracy and precision. Patient Plasma Analysis of VLCFAs. In order to test the clinical applicability of our assay, plasma samples collected from four subjects with X-linked adrenoleukodystrophy (ALD) and four normal controls were analyzed by MWCNT-based enrichment and SALDI-TOFMS. The SALDI-TOFMS spectra of VLCFAs in ALD patients and in normal controls are shown in Figure 4. The intensity ratio of C24:0/C22:0 was significantly higher in the ALD patients (1.02-1.49) than in the normal controls (0.64-0.76). The peaks at m/z 440.4 and 452.4 were assumed to be C23:0 and C24:1. 25,27,30,41 Table 2 summarizes the quantification data for VLCFAs concentrations and ratios in this study and compares them with the reference range reported by Schutgens. 27 In their study, samples from 109 healthy subjects and 41 patients with X-ALD were analyzed by GC-MS following derivatization by methyl ester and purification by thin-layer chromatography. The quantification results in our study were within the reported range. Usually, concentration ratios of C24:0/C22:0 are used to diagnose ALD. Stable-isotope quantification is important in the MALDI- TOFMS method because of the suppression effect of matrix interference peaks. In the MWCNT-based SALDI-TOFMS method described herein, the relative quantification can also be directly determined by measuring the ion peak intensities without the isotope-labeled internal standards calibration. Although the sample numbers are small, the concentration ratios of C24:0/C22:0 correlated well with the ion peak intensity ratios of C24:0/C22:0 (Pearson Correlation coefficient ) 0.88) (Figure S-8 in the (38) Koulman, A.; Petras, D.; Narayana, V. K.; Wang, L.; Volmer, D. A. Anal. Chem. 2009, 81, 7544–7551. (39) Sleno, L.; Volmer, D. A. Rapid Commun. Mass Spectrom. 2005, 19, 1928– 1936. (40) Hsu, W. Y.; Lo, W. Y.; Lai, C. C.; Tsai, F. J.; Tsai, C. H.; Tsai, Y.; Lin, W. D.; Chao, M. C. Rapid Commun. Mass Spectrom. 2007, 21, 1915–1919. (41) Aveldano, M. I.; Donnari, D. Clin. Chem. 1996, 42, 454–461. <strong>Analytical</strong> <strong>Chemistry</strong>, Vol. 82, No. 16, August 15, 2010 6819
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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
Page 23 and 24: Figure 1. GC/MS total ion chromatog
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Page 27 and 28: on a substrate are preferred. 20-24
Page 29 and 30: Figure 4. SERS analysis of NAADP co
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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 and 42: Figure 1. TEM images of the prepare
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Page 45 and 46: esult in a big SPR signal change wi
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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
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Page 59 and 60: Figure 4. (A) IRMS mass-44 chromato
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Page 65 and 66: Polymerase (1 U per sample). Reacti
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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
Page 93 and 94: Figure 5. Extinction spectra and co
Page 95 and 96: wished to explore the dehydration o
Page 97 and 98: constant medium for separation; we
Page 99 and 100: Figure 2. Standards of (Pi)n, n ) 1
Page 101 and 102: Figure 5. Quantitative calibration
Page 103 and 104: Anal. Chem. 2010, 82, 6847-6853 Met
Page 105 and 106: Figure 1. 226 Ra spectrum by liquid
Page 107 and 108: Table 1. Counting Properties and De
Page 109 and 110: Table 3. Analysis of 226 Ra in Sedi
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Page 115 and 116: Figure 5. Volcano plots detailing t
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Page 123 and 124: 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),
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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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Analytical Chemistry Chemical Cytometry Quantitates Superoxide

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