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ABSTRACT: A rapid method to determine drug resistance in bacteria based on mass spectrometry is presented. In it, a mass spectrum of an intact microorganism grown in drug-containing stable isotope-labeled media is compared with a mass spectrum of the intact microorganism grown in non-labeled media without the drug present. Drug resistance is determined by predicting characteristic mass shifts of one or more microorganism biomarkers using bioinformatics algorithms. Observing such characteristic mass shifts indicates that the microorganism is viable even in the presence of the drug, thus incorporating the isotopic label into characteristic biomarker molecules. The performance of the method is illustrated on the example of intact E. coli, grown in control (unlabeled) and (13)C-labeled media, and analyzed by MALDI TOF MS. Algorithms for data analysis are presented as well.
Journal of the American Society for Mass Spectrometry 04/2013; · 4.00 Impact Factor
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ABSTRACT: The capability to rapidly and confidently determine or confirm the sequences of short oligonucleotides, including native and chemically-modified DNA and RNA, is important for a number of fields. While matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) has been used previously to sequence short oligonucleotides, the typically low fragmentation efficiency of in-source or post-source decay processes necessitates the accumulation of a large number of spectra, thus limiting the throughput of these methods. Here we introduce a novel matrix, 1,5-diaminonapthalene (DAN), for facile in-source decay (ISD) of DNA and RNA molecular anions, which allows for rapid sequence confirmation. d-, w-, and y-series ions are prominent in the spectra, complementary to the (a-B)- and w- ions that are typically produced by MALDI post-source decay (PSD). Results are shown for several model DNA and RNA oligonucleotides, including combinations of DAN-induced fragmentation with true tandem TOF MS (MS/MS) for pseudo-MS(3) and "activated-ion PSD."
Journal of the American Society for Mass Spectrometry 02/2012; 23(4):773-7. · 4.00 Impact Factor
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ABSTRACT: We apply MALDI-TOF/TOF mass spectrometry for the rapid and high-confidence identification of intact Bacillus spore species. In this method, fragment ion spectra of whole (undigested) protein biomarkers are obtained without the need for biomarker prefractionation, digestion, separation, and cleanup. Laser-induced dissociation (unimolecular decay) of higher mass (>5 kDa) precursor ions in the first TOF analyzer is followed by reacceleration and subsequent high-resolution mass analysis of the resulting sequence-specific fragments in a reflectron TOF analyzer. In-house-developed software compares an experimental MS/MS spectrum with in silico-generated tandem mass spectra from all protein sequences, contained in a proteome database, with masses within a preset range around the precursor ion mass. A p-value, the probability that the observed matches between experimental and in silico-generated fragments occur by chance, is computed and used to rank the database proteins to identify the most plausible precursor protein. By inference, the source microorganism is then identified on the basis of the identification of individual, unique protein biomarker(s). As an example, intact Bacillus atrophaeus and Bacillus cereus spores, either pure or in mixtures, were unambiguously identified by this method after fragmenting and identifying individual small, acid-soluble spore proteins that are specific for each species. Factors such as experimental mass accuracy and number of detected fragment ions, precursor ion charge state, and sequence-specific fragmentation have been evaluated with the objective of extending the approach to other microorganisms. MALDI-TOF/TOF-MS in a lab setting is an efficient tool for in situ confirmation/verification of initial microorganism identification.
Analytical Chemistry 11/2005; 77(22):7455-61. · 5.86 Impact Factor
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ABSTRACT: Detection of Plasmodium falciparum malaria during pregnancy is complicated by sequestration of parasites in the placenta, which reduces peripheral blood microscopic detection. Laser desorption mass spectrometry (LDMS) has previously demonstrated sensitive detection of hemozoin from P. falciparum blood cultures and the ability to track parasitemia in a Plasmodium yoelii malaria mouse model. Here we use a simple, dilution in water, blood sample preparation protocol for LDMS detection of malaria in 45 asymptomatic, pregnant Zambian women. We compare LDMS to microscopy and polymerase chain reaction (PCR) analysis. All women were microscopy negative. LDMS detected P. falciparum hemozoin in 15 out of 45 women, while PCR results were positive in 25 women. Compared with PCR, which analyzed 20-30 microL of blood, the sensitivity of LDMS, which analyzed < 1 microL of blood, was 52%, with a specificity of 92%. LDMS is a potentially rapid and more sensitive alternate diagnostic method than microscopy.
The American journal of tropical medicine and hygiene 09/2005; 73(3):485-90. · 2.59 Impact Factor
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ABSTRACT: n the post-9/11 era, the asymmetric threats posed by terrorists or rogue states have created new challenges for the enhanced and efficient defense of the nation. For defense against chemical and biological weapons (cbW), integrated, multitiered, and "net-centric" systems are envisioned that will enable the rapid and cost-effective detection, confirmation, and response to a cbW attack. realization of this vision requires advances in the science and technology of chemical and biological sensor systems and multisource information fusion. our evolving counter-cbW capability has broader benefits to society, where, for example, new tools will become available to manage outbreaks of emerging natural infectious diseases or industrial accidents. here we highlight several key technolo-gies and the challenges pursued in support of this vision.
Johns Hopkins Apl Technical Digest 01/2005; 26. · 0.06 Impact Factor
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ABSTRACT: Rapid diagnosis leading to effective treatment is essential to control escalating infectious diseases such as malaria. Malaria pigment (hemozoin) detection by laser desorption mass spectometry (LDMS) was recently shown to be a sensitive (<10 parasites/muL) technique for detecting Plasmodium falciparum parasites cultured in human blood. To examine the use of LDMS in a rapid new malaria screening assay, we followed the time course of P. yoelii infections in mice in parallel with light microscopy and a colorimetric hemozoin assay. Hemozoin was detected by LDMS in 0.3 muL of blood within two days of infection independently of the inoculating dose of 10(6), 10(4), or 10(2) parasite-infected erythrocytes. Microscopy and colorimetric hemozoin determinations lagged the LDMS detection of infections by 2-4 and 3-5 days, respectively, except at the highest inoculation dose. The LDMS detection of hemozoin is a potentially more rapid screen than light microscopy for detecting malaria infection in this mouse model at parasitemias <0.1%.
The American journal of tropical medicine and hygiene 11/2004; 71(5):546-51. · 2.59 Impact Factor
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ABSTRACT: While radio frequency (RF) catheter ablation (RCA) procedures for treating ventricular arrhythmias have evolved significantly over the past several years, the use of RCA has been limited to treating slow ventricular tachycardias (VTs). In this paper, we present preliminary results from computer and animal studies to evaluate the accuracy of an algorithm that uses the single equivalent moving dipole (SEMD) model in an infinite homogeneous volume conductor to guide the RF catheter to the site of origin of the arrhythmia. Our method involves measuring body surface electrocardiographic (ECG) signals generated by arrhythmic activity and by bipolar current pulses emanating from a catheter tip, and representing each of them by a SEMD model source at each instant of the cardiac cycle, thus enabling rapid repositioning of the catheter tip requiring only a few cycles of the arrhythmia. We found that the SEMD model accurately reproduced body surface ECG signals with a correlation coefficients > 0.95. We used a variety of methods to estimate the uncertainty of the SEMD parameters due to measurement noise and found that at the time when the arrhythmia is mostly localized during the cardiac cycle, the estimates of the uncertainty of the spatial SEMD parameters (from ECG signals) are between 1 and 3 mm. We used pacing data from spatially separated epicardial sites in a swine model as surrogates for focal ventricular arrhythmic sources and found that the spatial SEMD estimates of the two pacing sites agreed with both their physical separation and orientation with respect to each other. In conclusion, our algorithm to estimate the SEMD parameters from body surface ECG can potentially be a useful method for rapidly positioning the catheter tip to the arrhythmic focus during an RCA procedure.
IEEE Transactions on Biomedical Engineering 12/2003; 50(12):1360-70. · 2.28 Impact Factor
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ABSTRACT: An improved data analysis method is described for rapid identification of intact microorganisms from MALDI-TOF-MS data. The method makes no use of mass spectral fingerprints. Instead, a microorganism database is automatically generated that contains biomarker masses derived from ribosomal protein sequences and a model of N-terminal Met loss. We quantitatively validate the method via a blind study that seeks to identify microorganisms with known ribosomal protein sequences. We also include in the database microorganisms with incompletely known sets of ribosomal proteins to test the specificity of the method. With an optimal MALDI protocol, and at the 95% confidence level, microorganisms represented in the database with 20 or more biomarkers (i.e., those with complete or nearly completely sequenced genomes) are correctly identified from their spectra 100% of the time, with no incorrect identifications. Microorganisms with seven or less biomarkers (i.e., incompletely sequenced genomes) are either not identified or misidentified. Robustness with respect to variations in sample preparation protocol and mass analysis protocol is demonstrated by collecting data with two different matrixes and under two different ion-mode configurations. Statistical analysis suggests that, even without further improvement, the method described here would successfully scale up to microorganism databases with roughly 1000 microorganisms. The results demonstrate that microorganism identification based on proteome data and modeling can perform as well as methods based on mass spectral fingerprinting.
Analytical Chemistry 09/2003; 75(15):3817-22. · 5.86 Impact Factor
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ABSTRACT: We propose a new method for guiding catheter ablation procedures to abolish sites of origin of arrhythmias. This method models both cardiac electrical activity and current pulses delivered from the tip of the ablation catheter with a single equivalent moving dipole (SEMD). The SEMD parameters are obtained from analysis of body surface potentials. In this paper we examine the feasibility of this method by evaluating the performance of an inverse algorithm we developed to localize the SEMD from the surface potentials. In computer simulations realistic levels of measurement noise led to uncertainties in SEMD location approximately 0.005 cm. Dipole orientation randomization contributed to increased uncertainty (0.04 cm) in SEMD location only when boundary effects were included. In ventricular pacing swine studies, we found that the SEMD model accurately accounted for electrocardiographic wave forms and that measurement noise led to an uncertainty of approximately 0.04 cm in the SEMD at 15 ms after the pacing spike. We have also found that the algorithm we developed to identify the SEMD parameters yielded positions for two spatially separated pacing sites that maintained their direction and were very close to their physical separation. These results suggest that the SEMD method may potentially be used to guide radio-frequency ablation procedures.
Annals of Biomedical Engineering 06/2003; 31(5):564-76. · 2.37 Impact Factor
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ABSTRACT: A novel method for acquisition and numerical analysis of matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectral data is described. The digitized ion current transient from each consecutive laser shot is first acquired and stored independently. Subsequently, statistical correlation parameters between all stored transients are computed. We illustrate the uses of this event-by-event analysis method for studies of sample surface heterogeneity as well as for elucidating the mechanisms of ion formation in MALDI. Other potential applications of the method are also outlined.
Rapid Communications in Mass Spectrometry 02/2003; 17(9):991-5. · 2.79 Impact Factor
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ABSTRACT: A quantitatively-reliable computer model of the human heart could be a very useful tool for the analysis and development
of potential tachyarrhythmia therapies. In this article, we present a new discrete cellular automata (CA) model of the human
ventricle that allows representation of important quantitative aspects of cardiac excitation waves. We use this model to analyze
a hypothetical technique for preemptive antitachycardia pacing using spatially-uniform, low-energy shocks. Because of the
computational efficiency and relative simplicity of the CA model, we can perform rapid simulations of a large number of pacing
interventions on a statistical ensemble of two-dimensional ventricular substrates with topology and dimensions similar to
the human ventricular epicardial surface. This allows us to obtain a good statistical profile of the intervention‘s efficacy
and proarrhythmic potential. Our study results suggest that while spatially-uniform, low-energy preemptive pacing shocks are
frequently successful at preventing tachyarrhythmia, they are often proarrhythmic. The simplicity of our CA model structure
and the clear physiological interpretation of its parameters allows us to examine the mechanisms of arrhythmia prevention
and spurious induction by the pacing shocks in terms of the generic properties of the waves. We believe such an approach can
potentially be very useful for the preliminary analysis of a wide variety of antiarrhythmic interventions, both electrical
and pharmacological.
Herzschrittmachertherapie & Elektrophysiologie 04/1999; 10(2):105-118.
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ABSTRACT: We present a new computer simulation technology suitable for rapid and quantitatively reliable simulation of propagating
excitation waves in anisotropic myocardium. Our model utilizes a finite element or cellular automata (CA) approach in which
the elements undergo transitions between a finite number of states (e.g., excited, refractory) according to specific rules.
The transition parameter values for the CA elements at each location are computed using the characteristic relations governing
propagation at the given point in the tissue, such as the anisotropy ratio and the dependence of the plane wave speed on diastolic
interval. The model is well-suited for analysis of arrhythmogenesis and hypothetical therapeutic interventions. Once the effects
of an antiarrhythmic drug or disease process on the characteristic relationships have been determined for different cardiac
cell types, the electrical activity in tissue with the modified properties can be simulated by our model. In this article,
we discuss the basic structure of the model and use it to demonstrate wavelet formation in myocardium with a fixed scar (infarct)
in the presence of a sodium channel blocker. This mechanism may help explain the proarrhythmic effects of these agents.
Herzschrittmachertherapie & Elektrophysiologie 04/1999; 10(2):92-104.
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Herzschrittmachertherapie & Elektrophysiologie 04/1999; 10(2):61-66.
