Fluorescence Imaging for Visualization of the Ion Cloud in a Quadrupole Ion Trap Mass Spectrometer

Laser-induced fluorescence is used to visualize populations of gaseous ions stored in a quadrupole ion trap (QIT) mass spectrometer. Presented images include the first fluorescence image of molecular ions collected under conditions typically used in mass spectrometry experiments. Under these “normal” mass spectrometry conditions, the radial (r) and axial (z) full-width at half maxima (FWHM) of the detected ion cloud are 615 and 214 μm, respectively, corresponding to ~6 % of r ₀ and ~3 % of z ₀ for the QIT used. The effects on the shape and size of the ion cloud caused by varying the pressure of helium bath gas, the number of trapped ions, and the Mathieu parameter q _z are visualized and discussed. When a “tickle voltage” is applied to the exit end-cap electrode, as is done in collisionally activated dissociation, a significant elongation in the axial, but not the radial, dimension of the ion cloud is apparent. Finally, using spectroscopically distinguishable fluorophores of two different m/z values, images are presented that illustrate stratification of the ion cloud; ions of lower m/z (higher q _z ) are located in the center of the trapping region, effectively excluding higher m/z (lower q _z ) ions, which form a surrounding layer. Fluorescence images such as those presented here provide a useful reference for better understanding the collective behavior of ions in radio frequency (rf) trapping devices and how phenomena such as collisions and space-charge affect ion distribution.

Quantitative,In Situ Visualization of Metal‐Ion Dissolution and Transport Using 1H Magnetic Resonance Imaging

Quantitative mapping of metal ions freely diffusing in solution is important across a diverse range of disciplines and is particularly significant for dissolution processes in batteries, metal corrosion, and electroplating/polishing of manufactured components. However, most current techniques are invasive, requiring sample extraction, insertion of an electrode, application of an electric potential or the inclusion of a molecular sensor. Thus, there is a need for techniques to visualize the distribution of metal ions non‐invasively, in situ, quantitatively, in three dimensions (3D) and in real time. Here we have used ¹H magnetic resonance imaging (MRI) to make quantitative 3D maps showing evolution of the distribution of Cu²⁺ ions, not directly visible by MRI, during the electrodissolution of copper, with high sensitivity and spatial resolution. The images are sensitive to the speciation of copper, the depletion of dissolved O₂ in the electrolyte and show the dissolution of Cu²⁺ ions is not uniform across the anode.     

Quantitative,In Situ Visualization of Metal‐Ion Dissolution and Transport Using 1H Magnetic Resonance Imaging

Quantitative mapping of metal ions freely diffusing in solution is important across a diverse range of disciplines and is particularly significant for dissolution processes in batteries, metal corrosion, and electroplating/polishing of manufactured components. However, most current techniques are invasive, requiring sample extraction, insertion of an electrode, application of an electric potential or the inclusion of a molecular sensor. Thus, there is a need for techniques to visualize the distribution of metal ions non‐invasively, in situ, quantitatively, in three dimensions (3D) and in real time. Here we have used ¹H magnetic resonance imaging (MRI) to make quantitative 3D maps showing evolution of the distribution of Cu²⁺ ions, not directly visible by MRI, during the electrodissolution of copper, with high sensitivity and spatial resolution. The images are sensitive to the speciation of copper, the depletion of dissolved O₂ in the electrolyte and show the dissolution of Cu²⁺ ions is not uniform across the anode.     

Determination of Ion Frequencies in a Quadrupole Ion Trap By Using a Fast Direct Current Pulse as Pump and a Laser Probe

A new technique has been developed which allows the direct measurement of frequencies of ions trapped in a quadrupole ion trap mass spectrometer. This pump/probe method employs a fast direct current (DC) pulse (pump) to displace a kinetically cooled ion population from the center of the trap, and a laser (probe) which recognizes when ions reappear at the center of the trap by the formation of photodissociation fragments. The translationally excited ions undergo periodic motion within the confines of the ion trap, and this periodic motion can be followed by recording the intensity of the photodissociation fragment as a function of the delay time between the DC pump and the laser probe. The DC pulse has a rise time of 15 ns; data are taken 1 ms after its application to allow stable ion motion to be sampled. Sampling of the ion cloud is done at 50 ns intervals, and fast Fourier transformation of the time-based data yields the ion frequencies and their relative magnitudes. Data are reported for ions derived from acetophenone (m/z 105) and 1,4-cyclohexadiene (m/z 80) under various trapping conditions corresponding to different Mathieu q_z values. The measured fundamental secular frequencies, f_z and f_r, are found to agree well with those predicted. The presence of higher order multipole contributions to the trapping field is evident from such ion frequencies as the drive frequency, f_RF,. The ability to measure ion frequencies under operating conditions provides a new tool for comparing simulated and experimental data. Simulation data from the program ITSIM, modified to account for the effects of collisions, are shown to predict the major frequency components observed in the experimental data.     

Biomolecular Imaging with a C60-SIMS/MALDI Dual Ion Source Hybrid Mass Spectrometer: Instrumentation,Matrix Enhancement,and Single Cell Analysis

We describe a hybrid MALDI/C₆₀-SIMS Q-TOF mass spectrometer and corresponding sample preparation protocols to image intact biomolecules and their fragments in mammalian spinal cord, individual invertebrate neurons, and cultured neuronal networks. A lateral spatial resolution of 10 μm was demonstrated, with further improvement feasible to 1 μm, sufficient to resolve cell outgrowth and interconnections in neuronal networks. The high mass resolution (>13,000 FWHM) and tandem mass spectrometry capability of this hybrid instrument enabled the confident identification of cellular metabolites. Sublimation of a suitable matrix, 2,5-dihydroxybenzoic acid, significantly enhanced the ion signal intensity for intact glycerophospholipid ions from mammalian nervous tissue, facilitating the acquisition of high-quality ion images for low-abundance biomolecules. These results illustrate that the combination of C₆₀-SIMS and MALDI mass spectrometry offers particular benefits for studies that require the imaging of intact biomolecules with high spatial and mass resolution, such as investigations of single cells, subcellular organelles, and communities of cells. Graphical Abstract

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Absolute Quantification of Rifampicin by MALDI Imaging Mass Spectrometry Using Multiple TOF/TOF Events in a Single Laser Shot

Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) allows for the visualization of molecular distributions within tissue sections. While providing excellent molecular specificity and spatial information, absolute quantification by MALDI IMS remains challenging. Especially in the low molecular weight region of the spectrum, analysis is complicated by matrix interferences and ionization suppression. Though tandem mass spectrometry (MS/MS) can be used to ensure chemical specificity and improve sensitivity by eliminating chemical noise, typical MALDI MS/MS modalities only scan for a single MS/MS event per laser shot. Herein, we describe TOF/TOF instrumentation that enables multiple fragmentation events to be performed in a single laser shot, allowing the intensity of the analyte to be referenced to the intensity of the internal standard in each laser shot while maintaining the benefits of MS/MS. This approach is illustrated by the quantitative analyses of rifampicin (RIF), an antibiotic used to treat tuberculosis, in pooled human plasma using rifapentine (RPT) as an internal standard. The results show greater than 4-fold improvements in relative standard deviation as well as improved coefficients of determination (R²) and accuracy (>93% quality controls, <9% relative errors). This technology is used as an imaging modality to measure absolute RIF concentrations in liver tissue from an animal dosed in vivo. Each microspot in the quantitative image measures the local RIF concentration in the tissue section, providing absolute pixel-to-pixel quantification from different tissue microenvironments. The average concentration determined by IMS is in agreement with the concentration determined by HPLC-MS/MS, showing a percent difference of 10.6%.

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Graphical Abstract ?

     

Degree of Ionization in MALDI of Peptides: Thermal Explanation for the Gas-Phase Ion Formation

Degree of ionization (DI) in matrix-assisted laser desorption ionization (MALDI) was measured for five peptides using α-cyano-4-hydroxycinnanmic acid (CHCA) as the matrix. DIs were low 10(-4) for peptides and 10(-7) for CHCA. Total number of ions (i.e., peptide plus matrix) was the same regardless of peptides and their concentration, setting the number of gas-phase ions generated from a pure matrix as the upper limit to that of peptide ions. Positively charged cluster ions were too weak to support the ion formation via such ions. The total number of gas-phase ions generated by MALDI, and that from pure CHCA, was unaffected by the laser pulse energy, invalidating laser-induced ionization of matrix molecules as the mechanism for the primary ion formation. Instead, the excitation of matrix by laser is simply a way of supplying thermal energy to the sample. Accepting strong Coulomb attraction felt by cations in a solid sample, we propose three hypotheses for gas-phase peptide ion formation. In Hypothesis 1, they originate from the dielectrically screened peptide ions in the sample. In Hypothesis 2, the preformed peptide ions are released as part of neutral ion pairs, which generate gas-phase peptide ions via reaction with matrix-derived cations. In Hypothesis 3, neutral peptides released by ablation get protonated via reaction with matrix-derived cations.     

Direct detection of peptides and proteins on a microfluidic platform with MALDI mass spectrometry

The ability to detect and quantify proteins of individual cells in high throughput is of enormous biological and clinical relevance. Most methods currently in use either require the measurement of large cell populations or are limited to the investigation of few cells at a time. In this report, we present the combination of a polydimethylsiloxane-based microfluidic device to a matrix-assisted laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF-MS) that allows the detection of as few as 300 molecules at the peptide level and ～10(6) to 10(7) molecules at the protein level. Moreover, we performed an immunoassay with subsequent MALDI-TOF-MS to capture and detect insulin immobilized on a surface (～0.05?mm(2)) in this device with a detection limit of 10(6) insulin molecules. This microfluidic-based approach therefore begins to approach the sample handling and sensitivity requirements for MS-based single-cell analysis of proteins and peptides and holds the potential for easy parallelization of immunoassays and other highly sensitive protein analyses.     

便携式检测仪检测呋喃丹和对氧磷的研究

基于胆碱酯酶被抑制前后反应速率不同的原理,研制了便携式荧光探针农药残留检测仪,并对检测条件进行了优化,获得呋喃丹和对氧磷的检出限分别为3．0和12μg／L,线性范围分别为3．0—37．8μg／L和12-350μg／L,该检测仪具有操作简便,检测怏速,可用于现场检测等优点,在对蔬菜和水果样品检测中,获得呋喃丹的回收率在87％～114％之间。     

MALDI Mass Spectrometry Imaging for the Distinction of Adenocarcinomas of the Pancreas and Biliary Tree

Pancreatic ductal adenocarcinoma and cholangiocarcinoma constitute two aggressive tumor types that originate from the epithelial lining of the excretory ducts of the pancreatobiliary tract. Given their close histomorphological resemblance, a correct diagnosis can be challenging and almost impossible without clinical information. In this study, we investigated whether mass spectrometric peptide features could be employed to distinguish pancreatic ductal adenocarcinoma from cholangiocarcinoma. Three tissue microarrays of formalin-fixed and paraffin-embedded material (FFPE) comprising 41 cases of pancreatic ductal adenocarcinoma and 41 cases of cholangiocarcinoma were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). The derived peptide features and respective intensities were used to build different supervised classification algorithms: gradient boosting (GB), support vector machine (SVM), and k-nearest neighbors (KNN). On a pixel-by-pixel level, a classification accuracy of up to 95% could be achieved. The tentative identification of discriminative tryptic peptide signatures revealed proteins that are involved in the epigenetic regulation of the genome and tumor microenvironment. Despite their histomorphological similarities, mass spectrometry imaging represents an efficient and reliable approach for the distinction of PDAC from CC, offering a promising complementary or alternative approach to the existing tools used in diagnostics such as immunohistochemistry.     

Ion yields of thin MALDI samples: dependence on matrix and metal substrate and implications for models

Thin MALDI samples can perform differently than thicker samples, on metal substrates. Divergent results and models for the effect have been presented. Positive and negative yields are investigated here for three matrixes (2,5-dihydroxybenzoic acid (DHB), sinapinic acid, and alpha-cyano 4-hydroxycinnamic acid) on stainless steel and gold substrates. Samples were electrosprayed for uniformity and thickness control and imaged across a metal-metal boundary. Thin sample enhancement is found in both polarities for all three matrixes on a steel substrate. On gold, only alpha-cyano-4-hydroxycinnamic acid shows enhancement. These and earlier data are used to evaluate two models. The first is based on one-photon photoelectron emission from the metal; the second one, on two-photon matrix ionization at the metal interface. The surface-enhanced matrix photoionization model best fits the evidence, including the fluence dependence of electron emission from DHB on steel.     

Functional Hybrid Molecules for the Visualization of Cancer: PESIN-Homodimers Combined with Multimodal Molecular Imaging Probes for Positron Emission Tomography and Optical Imaging: Suited for Tracking of GRPR-Positive Malignant Tissue**

We describe multimodal imaging probes for gastrin-releasing peptide receptor (GRPR)-specific targeting suited for positron emission tomography and optical imaging (PET/OI), consisting of PESIN (PEG₃-BBN_7-14) dimers connected to multimodal imaging subunits. These multimodal agents comprise a fluorescent dye for OI and the chelator ((1,4,7-triazacyclononane-4,7-diyl)diacetic acid-1-glutaric acid) (NODA-GA) for PET radiometal isotope labelling. Special focus was put on the influence of the used dyes on the properties of the whole bioconjugates. For this, several compounds with different fluorescent dyes and non-dye carrying subunits were synthesized and investigated. As fluorescent dyes, dansyl, NBD, derivatives of fluorescein, coumarin and rhodamine as well as three pyrilium-based dyes were employed. Considerable influence of the charge of the colored unit on hydrophilicity as well as in vitro target receptor binding was observed and classified. High radiochemical yields and purities were found during radiolabeling of the multimodal imaging subunits as well as their GRPR-specific bioconjugates with ⁶⁸Ga. Examinations of the photophysical properties of both molecule species displayed no loss or alteration of fluorescence characteristics.     

Implementation of a Gaussian Beam Laser and Aspheric Optics for High Spatial Resolution MALDI Imaging MS

We have investigated the use of a Gaussian beam laser for MALDI Imaging Mass Spectrometry to provide a precisely defined laser spot of 5 μm diameter on target using a commercial MALDI TOF instrument originally designed to produce a 20 μm diameter laser beam spot at its smallest setting. A Gaussian beam laser was installed in the instrument in combination with an aspheric focusing lens. This ion source produced sharp ion images at 5 μm spatial resolution with signals of high intensity as shown for images from thin tissue sections of mouse brain.

Highly Luminescent Crystalline Sponge: Sensing Properties and Direct X-ray Visualization of the Substrates

A phenomenon of crystalline sponge is represented by guest-dependent structural fluidity of the host polymeric lattice in highly crystalline sorbents, such as metal-organic frameworks, driven by multiple weak intermolecular interactions. Such induced fitting in MOFs is a valuable property in selective adsorption, guest determination by single-crystal XRD and in-situ structural analysis under external stimuli. In this work, a porous three-dimensional metal-organic framework [Eu₂(DMF)₄(ttdc)₃]·4.45DMF (1_DMF; DMF = N,N-dimethylformamide, ttdc^2– = trans-thienothiophenedicarboxylate anion) was applied as a crystalline sponge bearing luminescent functionality to couple its sensing properties with direct structural determination of the adsorbed molecules. As a result, the paper discusses crystal structures and luminescent properties for the successfully obtained new adducts with the crystallographic formulae [Eu₂(DMSO)₄(ttdc)₃]·2.5DMSO·2.2H₂O (1_DMSO; DMSO = dimethylsulfoxide), [Eu₂(DMF)₄(ttdc)₃]·3phet (1_phet; phet = phenylethanal) and [Eu₂(DMF)_3.5(cin)_0.5(ttdc)₃]·1.64cin (1_cin; cin = trans-cinnamaldehyde). As a result of inclusion of DMSO into 1, a slight increase in the quantum yield and excited state phosphorescence lifetime was observed, while the adsorption of phet leads to a considerable (up to three times) decrease in the corresponding values. The incorporation of cinnamal results in a full quenching of QY, from 20% down to zero, and a more than order of magnitude diminishing of the excited state lifetime compared to the initial 1_DMF. The effective sensing of cinnamal was explained from the structural point of view by its direct coordination to the Eu³⁺ emitter, as well as by multiple weak intermolecular interactions with ttdc antenna ligand, both capable of enhancing the non-radiative energy dissipation.     

高效液相色谱－荧光检测法测定蒽和荧蒽

采用高效液相色谱－荧光检测法测定多环芳烃化合物蒽和荧蒽。在Ｓｈｉｍ－ｐａｃｋＣＬＣ－ｐｈｅｎｙｌ柱上，以５０％的乙腈水溶液作流动相，１２ｍｉｎ内实现了两组分的同时分离测定。线性范围为０～８０μｇ／ｍＬ，检测限低于５０ｎｇ／ｍＬ，相对标准偏差小于１％。     

Use of an Imaging Spectrograph System with a Fiber Optical Sensor and Two Dimensional Cooled Ccd Detector For Solid Surface Room Temperature Phosphorescence of Pesticides

ABSTRACT

This paper presents a development of a fiber optic sensor for solid surface room temperature phosphorescence of pesticides. The system is based on an imaging spectrograph coupled with a cooled two-dimensional charge coupled device detector. An optimization approach using a different mode of reading out data, the effects of moisture in solid matrix phosphorescence and the effects of heavy atoms are discussed. Spectral characterizations on Whatman N⁰1 filter papers of Warfarin, Naptalam, and naphthalene-type pesticides were evaluated. Limits of detection at ng to pg levels were estimated for all compounds. Satisfactory reproducibility of measurements was observed. The standard addition method was used to determine concentrations of Warfarin and Naptalam pesticides present together in a mixture. Results obtained show the potential of the device for the quantitative analysis of environmental pollutants.     

Dual-Phase Thermally Activated Delayed Fluorescence Luminogens: A Material for Time-Resolved Imaging Independent of Probe Pretreatment and Probe Concentration

Developing luminescent probes with long lifetime and high emission efficiency is essential for time-resolved imaging. However, the practical applications usually suffer from emission quenching of traditional luminogens in aggregated states, or from weak emission of aggregation-induced emission type luminogens in monomeric states. Herein, we overcome this dilemma by a rigid-and-flexible alternation design in donor–acceptor–donor skeletons, to achieve a thermally activated delayed fluorescence luminogen with high emission efficiency both in the monomeric state (quantum yield up to 35.3 %) and in the aggregated state (quantum yield up to 30.8 %). Such a dual-phase strong and long-lived emission allows a time-resolved luminescence imaging, with an efficiency independent of probe pretreatment and probe concentration. The findings open opportunities for developing luminescent probes with a usage in larger temporal and spatial scales.     

Determination of Homologue Imidazolium Ionic Liquid Cations by Ion Chromatography Using a Carboxyl Acid Cation-Exchange Column with Direct Conductivity Detection

An investigation into the determination of four imidazolium ionic liquid cations by ion chromatography using a carboxyl acid cation exchange column and direct conductivity detection was carried out. This research has developed a simple, selective, and accurate ion chromatographic method for separation of imidazolium ionic liquid cations. Detection limits (S/N = 3) for the cations were 3.2–24.3 mg/L. Relative standard deviations (RSD, n = 5) for peak areas were less than 1.6%. The method has been successfully applied to the determination of two ionic liquids synthesized by organic chemistry lab.